1 00:00:00,500 --> 00:00:02,830 The following content is provided under a Creative 2 00:00:02,830 --> 00:00:04,370 Commons license. 3 00:00:04,370 --> 00:00:06,670 Your support will help MIT OpenCourseWare 4 00:00:06,670 --> 00:00:11,030 continue to offer high quality educational resources for free. 5 00:00:11,030 --> 00:00:13,660 To make a donation or view additional materials 6 00:00:13,660 --> 00:00:17,610 from hundreds of MIT courses, visit MIT OpenCourseWare 7 00:00:17,610 --> 00:00:18,520 at ocw.mit.edu. 8 00:00:25,710 --> 00:00:28,740 JOANNE STUBBE: What I want to do is sort of introduce you 9 00:00:28,740 --> 00:00:31,890 to the second half of the course, where we're going, 10 00:00:31,890 --> 00:00:33,850 and what topics we're going to be covering. 11 00:00:33,850 --> 00:00:36,600 And then I'll start in module 5. 12 00:00:36,600 --> 00:00:42,180 So as with the first half of the course, we have four modules. 13 00:00:42,180 --> 00:00:45,790 The first half the courses was pretty well organized-- 14 00:00:45,790 --> 00:00:47,910 that is, you went from here to here to here. 15 00:00:47,910 --> 00:00:49,140 It all sort of made sense. 16 00:00:49,140 --> 00:00:51,750 This half of the course doesn't do that. 17 00:00:51,750 --> 00:00:53,520 We're talking about-- there are hundreds 18 00:00:53,520 --> 00:00:57,660 of topics in biochemistry, and any one of them 19 00:00:57,660 --> 00:00:59,880 is exciting and important. 20 00:00:59,880 --> 00:01:03,300 And these are the ones we're talking about this semester. 21 00:01:03,300 --> 00:01:04,890 And again, the focus will be sort 22 00:01:04,890 --> 00:01:07,380 of trying to get you to think about, 23 00:01:07,380 --> 00:01:10,180 what is the evidence that supports 24 00:01:10,180 --> 00:01:12,310 the model I'm going to present. 25 00:01:12,310 --> 00:01:18,120 So in module 5, we're going to be talking about the terpenome. 26 00:01:18,120 --> 00:01:20,460 And I'm going to be talking about, most of you 27 00:01:20,460 --> 00:01:25,320 have seen in the last module, with polyketide synthases, 28 00:01:25,320 --> 00:01:27,840 you have aldol reactions and claison reactions 29 00:01:27,840 --> 00:01:29,940 to form carbon carbon bonds. 30 00:01:29,940 --> 00:01:33,510 In this module, you're going to be introduced to another way 31 00:01:33,510 --> 00:01:35,410 to form carbon carbon bonds. 32 00:01:35,410 --> 00:01:38,610 And that's through C5 units. 33 00:01:38,610 --> 00:01:43,110 And C5 units are the basis for forming cholesterol, which is 34 00:01:43,110 --> 00:01:46,520 really the focus of module 5. 35 00:01:46,520 --> 00:01:48,540 And what we're going to be talking about 36 00:01:48,540 --> 00:01:51,300 is initially cholesterol biosynthesis, 37 00:01:51,300 --> 00:01:53,910 that will be this lecture and probably 38 00:01:53,910 --> 00:01:56,190 most of the next lecture. 39 00:01:56,190 --> 00:01:59,370 And then you all know from eating McDonald's hamburgers, 40 00:01:59,370 --> 00:02:01,830 you get a lot of cholesterol in your diet. 41 00:02:01,830 --> 00:02:05,760 And the question is, how do you control cholesterol levels? 42 00:02:05,760 --> 00:02:08,550 And so this semester, the second part of the semester 43 00:02:08,550 --> 00:02:11,370 is really focus on the question of homeostasis. 44 00:02:11,370 --> 00:02:12,660 Cholesterol is essential. 45 00:02:12,660 --> 00:02:14,460 If you have too much you've got problems. 46 00:02:14,460 --> 00:02:17,130 Doesn't matter what pathway you're talking about, 47 00:02:17,130 --> 00:02:20,430 if you have too much or you have too little, you have problems. 48 00:02:20,430 --> 00:02:22,668 So how do you control everything? 49 00:02:22,668 --> 00:02:24,960 And we're going to be talking about cholesterol sensing 50 00:02:24,960 --> 00:02:26,760 and regulation. 51 00:02:26,760 --> 00:02:29,700 And we're going to come back to a topic you covered 52 00:02:29,700 --> 00:02:34,920 in the first half of the course with ClpX and ClpP protein 53 00:02:34,920 --> 00:02:36,660 mediated degradation, because it plays 54 00:02:36,660 --> 00:02:41,460 a central role in controlling cholesterol homeostasis. 55 00:02:41,460 --> 00:02:46,080 So that will be the first module, module 5. 56 00:02:46,080 --> 00:02:51,180 Module 6 is also going to be a module on homeostasis. 57 00:02:51,180 --> 00:02:53,580 It's my feeling that in introductory courses, 58 00:02:53,580 --> 00:02:57,300 people don't get introduced enough to metals. 59 00:02:57,300 --> 00:03:01,410 And 50% of all the proteins inside of us 60 00:03:01,410 --> 00:03:04,090 bind metals and do something with them. 61 00:03:04,090 --> 00:03:05,790 And so in this module, I'm going to talk 62 00:03:05,790 --> 00:03:07,680 about metal homeostasis. 63 00:03:07,680 --> 00:03:10,950 And I'm really going to focus on iron. 64 00:03:10,950 --> 00:03:12,750 And you'll see why I'm going to focus 65 00:03:12,750 --> 00:03:14,110 on iron when we get there. 66 00:03:14,110 --> 00:03:17,880 But we're going to talk about iron sensing and regulation, 67 00:03:17,880 --> 00:03:20,730 initially in humans, and then we're 68 00:03:20,730 --> 00:03:24,435 going to focus on the war between pathogenic organisms 69 00:03:24,435 --> 00:03:27,570 for iron and humans for iron, since iron 70 00:03:27,570 --> 00:03:33,000 is essential for almost all organisms to survive. 71 00:03:33,000 --> 00:03:35,850 The third module is sort of-- 72 00:03:35,850 --> 00:03:40,320 module 7-- sort of follows from module 6. 73 00:03:40,320 --> 00:03:45,780 And it's a topic that I've been following for 30 years, 74 00:03:45,780 --> 00:03:48,750 and it irritates the hell out of me. 75 00:03:48,750 --> 00:03:52,650 So everybody talks about reactive oxygen species, 76 00:03:52,650 --> 00:03:54,150 and how bad they are. 77 00:03:54,150 --> 00:03:56,020 You can't-- you can listen to NPR, 78 00:03:56,020 --> 00:03:58,320 you can read it in The New York Times. 79 00:03:58,320 --> 00:04:00,600 What are reactive oxygen species? 80 00:04:00,600 --> 00:04:02,700 As a chemist, what are they? 81 00:04:02,700 --> 00:04:06,930 So we'll define what reactive oxygen species are. 82 00:04:06,930 --> 00:04:09,010 Many of you know that they're bad. 83 00:04:09,010 --> 00:04:13,200 That's why you eat vitamin C and vitamin E. 84 00:04:13,200 --> 00:04:17,190 And they're involved, in fact, in defense 85 00:04:17,190 --> 00:04:19,410 against some microorganisms. 86 00:04:19,410 --> 00:04:21,279 But also they're good. 87 00:04:21,279 --> 00:04:23,850 They're now known to be involved in signaling. 88 00:04:23,850 --> 00:04:26,820 So again, it's a question of homeostasis. 89 00:04:26,820 --> 00:04:29,130 And so you need to understand the chemistry 90 00:04:29,130 --> 00:04:33,480 of what these species do, and where they can go astray, 91 00:04:33,480 --> 00:04:35,430 or where you can harness the reactivity 92 00:04:35,430 --> 00:04:37,690 to do something important. 93 00:04:37,690 --> 00:04:40,380 And the last section-- hopefully we'll get there this year, 94 00:04:40,380 --> 00:04:41,520 I'm trying my best-- 95 00:04:41,520 --> 00:04:44,080 this is the area I know most about, 96 00:04:44,080 --> 00:04:47,100 we're going to talk about nucleotide metabolism. 97 00:04:47,100 --> 00:04:50,640 In 5.07, they don't talk about nucleotide metabolism at all. 98 00:04:50,640 --> 00:04:52,710 And I would say in the next decade, 99 00:04:52,710 --> 00:04:56,220 you're going to see a lot about nucleotide metabolism. 100 00:04:56,220 --> 00:04:59,670 Where have you seen ATP and GTP in the first part 101 00:04:59,670 --> 00:05:00,560 of this course? 102 00:05:00,560 --> 00:05:01,990 Everywhere. 103 00:05:01,990 --> 00:05:03,720 How do we control all of that? 104 00:05:03,720 --> 00:05:05,070 Pretty important. 105 00:05:05,070 --> 00:05:07,170 What are the questions we're going to be asking? 106 00:05:07,170 --> 00:05:08,190 Where does it come from? 107 00:05:08,190 --> 00:05:10,080 And how do we control the levels, 108 00:05:10,080 --> 00:05:13,050 which is central to all of metabolism? 109 00:05:13,050 --> 00:05:15,570 Hopefully we'll get there and discuss that. 110 00:05:18,170 --> 00:05:24,110 So the required reading has now been posted on Stellar. 111 00:05:24,110 --> 00:05:28,330 And there are-- there is really sort of three 112 00:05:28,330 --> 00:05:29,330 things you need to read. 113 00:05:29,330 --> 00:05:32,872 One is about a short review on the terpenome, which 114 00:05:32,872 --> 00:05:34,580 is what I'm going to start talking about. 115 00:05:34,580 --> 00:05:38,270 The second is lipid metabolism that's 116 00:05:38,270 --> 00:05:40,400 been taken from Voet & Voet, you can 117 00:05:40,400 --> 00:05:44,795 go back in any basic textbook and read the section, 118 00:05:44,795 --> 00:05:46,670 because it's central to thinking about what's 119 00:05:46,670 --> 00:05:49,730 going on with cholesterol. 120 00:05:49,730 --> 00:05:52,520 And then you'll see that these are 121 00:05:52,520 --> 00:05:55,190 two of my favorite guys, Brown and Goldstein. 122 00:05:55,190 --> 00:05:57,050 They won the Nobel Prize for their work, 123 00:05:57,050 --> 00:06:01,310 but in reality they should have won at least two Nobel prizes 124 00:06:01,310 --> 00:06:02,120 for their work. 125 00:06:02,120 --> 00:06:05,210 I mean, you can never not listen to them talk 126 00:06:05,210 --> 00:06:06,650 and not get excited. 127 00:06:06,650 --> 00:06:08,540 I mean, they always have something 128 00:06:08,540 --> 00:06:11,330 important and exciting to say. 129 00:06:11,330 --> 00:06:13,890 So they-- last year we used this review. 130 00:06:13,890 --> 00:06:15,440 There's a new review. 131 00:06:15,440 --> 00:06:18,080 They're different, they're both pretty short. 132 00:06:18,080 --> 00:06:21,530 Pick which one you want, they cover the material. 133 00:06:21,530 --> 00:06:25,610 And then this one doesn't cover the most recent material 134 00:06:25,610 --> 00:06:26,200 as well. 135 00:06:26,200 --> 00:06:30,170 So here's another short review that covers that material. 136 00:06:30,170 --> 00:06:33,710 So these guys here will give you an overview of what 137 00:06:33,710 --> 00:06:35,932 I'm going to be talking about. 138 00:06:35,932 --> 00:06:40,310 And they've all been posted on the website already. 139 00:06:40,310 --> 00:06:43,730 So cholesterol homeostasis-- 140 00:06:43,730 --> 00:06:46,070 I never end a lecture on time. 141 00:06:46,070 --> 00:06:48,920 You'll find out that I'm trying my best, anyhow. 142 00:06:48,920 --> 00:06:52,050 We have about five lectures we're going to be covering. 143 00:06:52,050 --> 00:06:55,040 And this is where we're going. 144 00:06:55,040 --> 00:06:56,900 And the first one, today's lecture, we're 145 00:06:56,900 --> 00:06:59,660 going to be talking about a new way 146 00:06:59,660 --> 00:07:04,070 to form carbon carbon bonds through C5 units. 147 00:07:04,070 --> 00:07:09,920 And we'll be talking about the terpenome, where 148 00:07:09,920 --> 00:07:13,400 there are a huge number of natural products, 149 00:07:13,400 --> 00:07:15,470 distinct from polyketide synthases 150 00:07:15,470 --> 00:07:19,100 and non-ribosomal polypeptide synthases 151 00:07:19,100 --> 00:07:21,770 that you just finished talking about. 152 00:07:21,770 --> 00:07:26,570 We want to get to-- we'll see in the biosynthetic pathway 153 00:07:26,570 --> 00:07:30,590 to get to these C5 building blocks, 154 00:07:30,590 --> 00:07:34,880 we need to get to a metabolite called mevalonic acid, that's 155 00:07:34,880 --> 00:07:40,040 front and central in cholesterol biosynthesis. 156 00:07:40,040 --> 00:07:41,690 So we need to get that far. 157 00:07:41,690 --> 00:07:44,960 And then we need to get from mevalonic acid 158 00:07:44,960 --> 00:07:47,480 into cholesterol. 159 00:07:47,480 --> 00:07:50,420 And so we'll be talking about this the first couple 160 00:07:50,420 --> 00:07:52,620 of lectures. 161 00:07:52,620 --> 00:07:54,800 In addition to making cholesterol, 162 00:07:54,800 --> 00:07:57,410 you get a lot of cholesterol from your diet. 163 00:07:57,410 --> 00:08:00,440 And so the question is, how does it 164 00:08:00,440 --> 00:08:04,700 get from your diet, transferred through the plasma, 165 00:08:04,700 --> 00:08:06,500 and taken up into cells? 166 00:08:06,500 --> 00:08:10,430 And this section, we'll describe the discovery 167 00:08:10,430 --> 00:08:13,880 of receptor mediated endocytosis, 168 00:08:13,880 --> 00:08:16,490 by Brown and Goldstein, that's now known 169 00:08:16,490 --> 00:08:17,840 to be prevalent everywhere. 170 00:08:17,840 --> 00:08:22,130 So the module-- module 6, you take up 171 00:08:22,130 --> 00:08:26,900 iron by receptor mediated endocytosis. 172 00:08:26,900 --> 00:08:32,795 In module 7, growth factors are involved in receptor mediated 173 00:08:32,795 --> 00:08:34,059 endocytosis. 174 00:08:34,059 --> 00:08:37,580 So it represents a general paradigm 175 00:08:37,580 --> 00:08:41,330 that happens all the time over and over again in biology. 176 00:08:41,330 --> 00:08:43,789 And then we're going to ask the question, 177 00:08:43,789 --> 00:08:45,590 how do you sense the cholesterol? 178 00:08:45,590 --> 00:08:49,220 And we're going to be doing two recitations on this. 179 00:08:49,220 --> 00:08:53,600 And then we'll have a few lectures on the machinery 180 00:08:53,600 --> 00:08:58,460 that sense cholesterol sterile responsive binding proteins, 181 00:08:58,460 --> 00:09:02,210 in a molecule called SCAP, and another molecule-- 182 00:09:02,210 --> 00:09:05,750 both of these are proteins-- called INSIGs. 183 00:09:05,750 --> 00:09:08,120 Everything is located in a membrane-- that's something 184 00:09:08,120 --> 00:09:10,190 you haven't been exposed to. 185 00:09:10,190 --> 00:09:12,110 How do you control everything when you 186 00:09:12,110 --> 00:09:14,792 have stuff stuck in a membrane. 187 00:09:14,792 --> 00:09:16,250 And then we'll come back at the end 188 00:09:16,250 --> 00:09:20,810 to look at the rate limiting step 189 00:09:20,810 --> 00:09:26,480 in formation of mevalonic acid, HMGCoA reductase, 190 00:09:26,480 --> 00:09:29,900 and how that plays a key role, since it's 191 00:09:29,900 --> 00:09:32,180 involved in making cholesterol. 192 00:09:32,180 --> 00:09:34,970 How do we control and regulate that protein, also 193 00:09:34,970 --> 00:09:37,160 in the ER membrane. 194 00:09:37,160 --> 00:09:41,390 And we'll see it requires ubiquitin mediated protein 195 00:09:41,390 --> 00:09:42,150 degradation. 196 00:09:42,150 --> 00:09:44,330 And so that's why I'm going to come back, 197 00:09:44,330 --> 00:09:46,520 and we're going to spend a little bit of time 198 00:09:46,520 --> 00:09:49,880 talking about this process in eukaryotic systems. 199 00:09:49,880 --> 00:09:52,440 And actually, in all the other modules, 200 00:09:52,440 --> 00:09:55,880 you're going to see ubiquitin mediated protein degradation. 201 00:09:55,880 --> 00:10:01,520 And finally, this week in recitation, there's a new-- 202 00:10:01,520 --> 00:10:03,710 not new, it was discovered in 2003-- 203 00:10:03,710 --> 00:10:09,860 a new target for drug therapy in controlling cholesterol levels. 204 00:10:09,860 --> 00:10:13,340 And there was a paper published this year 205 00:10:13,340 --> 00:10:16,260 to show that this is, in fact, a good target. 206 00:10:16,260 --> 00:10:21,670 And this paper used CRISPR-Cas9 technology. 207 00:10:21,670 --> 00:10:22,780 So I'm going to-- 208 00:10:22,780 --> 00:10:25,400 even though I'm not an expert in that, my lab hasn't used it-- 209 00:10:25,400 --> 00:10:27,400 I'm going to introduce you to this technology, 210 00:10:27,400 --> 00:10:31,210 and then focus on why we think this is a good new target, 211 00:10:31,210 --> 00:10:33,010 and what the targeting is. 212 00:10:33,010 --> 00:10:35,890 So that's where we're going. 213 00:10:35,890 --> 00:10:39,250 So the terpenome-- let's see if I can 214 00:10:39,250 --> 00:10:40,750 remember what I'm going to say. 215 00:10:40,750 --> 00:10:44,750 The first thing I want to say is-- let me just get all this-- 216 00:10:44,750 --> 00:10:47,000 OK, the first thing I want to tell you something about 217 00:10:47,000 --> 00:10:48,110 is the nomenclature-- 218 00:10:52,250 --> 00:10:55,760 and all terpenes are either called 219 00:10:55,760 --> 00:11:07,510 isoprenoids or terpenoids, and they're all made from C5-- 220 00:11:07,510 --> 00:11:10,535 a C5 hydrocarbon skeleton. 221 00:11:14,040 --> 00:11:19,910 And this C5 hydrocarbon skeleton is an isoprene. 222 00:11:19,910 --> 00:11:22,490 So this is the key building block 223 00:11:22,490 --> 00:11:25,070 that you're going to see over and over again 224 00:11:25,070 --> 00:11:29,690 over the course of the first couple of lectures. 225 00:11:29,690 --> 00:11:34,355 So an isoprenoid is, in general, linear. 226 00:11:37,610 --> 00:11:40,640 And it's made of n of these C5 units. 227 00:11:40,640 --> 00:11:49,010 So n can be 2 to thousands. 228 00:11:49,010 --> 00:11:51,130 And I'll give you examples of these. 229 00:11:51,130 --> 00:11:56,040 And again-- and then the terpenoids-- 230 00:11:56,040 --> 00:12:00,110 so let's just use terpenoids over here-- 231 00:12:00,110 --> 00:12:05,360 in general are also made of C5-- 232 00:12:05,360 --> 00:12:08,420 C5 units. 233 00:12:08,420 --> 00:12:18,500 But often, they're oxidized, cyclized, 234 00:12:18,500 --> 00:12:20,100 and sometimes rearranged. 235 00:12:23,820 --> 00:12:26,850 So my goal today really sort of is introduce you 236 00:12:26,850 --> 00:12:30,990 to this huge class of natural products. 237 00:12:30,990 --> 00:12:32,550 And give you some examples of this, 238 00:12:32,550 --> 00:12:36,780 and then start focusing on how we get the building blocks. 239 00:12:36,780 --> 00:12:38,550 Do you think iso-- 240 00:12:38,550 --> 00:12:40,335 this isoprene can be a building block? 241 00:12:40,335 --> 00:12:42,060 It's chemically not very reactive. 242 00:12:42,060 --> 00:12:43,740 So no, we have to convert-- 243 00:12:43,740 --> 00:12:47,460 we have to convert this into the chemically reactive building 244 00:12:47,460 --> 00:12:48,390 block. 245 00:12:48,390 --> 00:12:54,500 So while isoprene gives you the C5 unit, 246 00:12:54,500 --> 00:12:59,235 our focus today is going to be on creating the building 247 00:12:59,235 --> 00:12:59,735 blocks. 248 00:13:09,550 --> 00:13:13,120 And again, the building blocks are going to be-- 249 00:13:13,120 --> 00:13:15,610 these are the guys you're going to see over and over again. 250 00:13:15,610 --> 00:13:20,310 1, 2, 3, 4, 5-- 251 00:13:20,310 --> 00:13:22,785 so does everybody know what PPI means, so I 252 00:13:22,785 --> 00:13:23,910 don't have to write it out? 253 00:13:23,910 --> 00:13:26,710 pyrophosphate-- we're going to see this over and over again. 254 00:13:26,710 --> 00:13:29,730 You've seen this in the first part of the semester. 255 00:13:29,730 --> 00:13:35,490 And this isopentenyl pyrophosphate, or IPP. 256 00:13:38,220 --> 00:13:39,750 And we're going to see this-- 257 00:13:39,750 --> 00:13:42,660 if you look at this this hydrogen, what's 258 00:13:42,660 --> 00:13:47,210 interesting about this hydrogen, chemically? 259 00:13:47,210 --> 00:13:50,600 What's the pKa of that hydrogen, compared 260 00:13:50,600 --> 00:13:52,895 to if it was this hydrogen? 261 00:13:55,630 --> 00:13:56,370 It's much lower. 262 00:13:56,370 --> 00:14:01,390 Yeah, so it can form allylic cations or anions. 263 00:14:01,390 --> 00:14:03,670 And so this can readily isomerize 264 00:14:03,670 --> 00:14:10,340 to form this species, which also plays, 265 00:14:10,340 --> 00:14:17,720 which is dimethylalyl pyrophosphate, which 266 00:14:17,720 --> 00:14:21,170 is the other key building block that we're 267 00:14:21,170 --> 00:14:22,140 going to be looking at. 268 00:14:22,140 --> 00:14:26,420 So currently, it's estimated from the latest paper 269 00:14:26,420 --> 00:14:30,170 that I've read that these are the building blocks for what 270 00:14:30,170 --> 00:14:31,550 we call the terpenome. 271 00:14:34,610 --> 00:14:39,290 And it's estimated that there are greater than 70,000 272 00:14:39,290 --> 00:14:40,055 natural products. 273 00:14:45,480 --> 00:14:47,340 Now in contrast to what you've learned 274 00:14:47,340 --> 00:14:50,620 about with non-ribosomal polypeptides 275 00:14:50,620 --> 00:14:55,710 synthases and polyketide synthases, 276 00:14:55,710 --> 00:14:58,500 where you sort of can find everything in an operon, 277 00:14:58,500 --> 00:15:01,830 and you can sort of understand how your molecules could 278 00:15:01,830 --> 00:15:03,030 be put together. 279 00:15:03,030 --> 00:15:04,830 It's not so trivial with terpenes. 280 00:15:04,830 --> 00:15:09,480 There is no such logic in these systems. 281 00:15:09,480 --> 00:15:14,280 So let's just look at what some of these molecules 282 00:15:14,280 --> 00:15:18,860 actually are so you know why they're important. 283 00:15:18,860 --> 00:15:26,980 And, OK, so so in the center of everything 284 00:15:26,980 --> 00:15:30,600 are these two guys, our two building blocks. 285 00:15:30,600 --> 00:15:39,810 And these building blocks can go to the fat soluble vitamins-- 286 00:15:39,810 --> 00:15:44,110 so for example A and K. So if you look over here, 287 00:15:44,110 --> 00:15:45,545 you have vitamin A-- 288 00:15:45,545 --> 00:15:47,020 I knew this was going to happen. 289 00:15:47,020 --> 00:15:50,820 My late-- my pointers are not working very well. 290 00:15:50,820 --> 00:15:54,390 I need-- OK, so anyhow you have vitamin A and you have vitamin 291 00:15:54,390 --> 00:15:55,860 K. And where do you see-- you can 292 00:15:55,860 --> 00:15:59,430 see here readily that you have these C5 units somehow 293 00:15:59,430 --> 00:16:00,110 stuck together. 294 00:16:00,110 --> 00:16:01,720 And you have to ask the question, 295 00:16:01,720 --> 00:16:04,120 where does the rest of this come from? 296 00:16:04,120 --> 00:16:06,840 So fat soluble vitamins, which we're not 297 00:16:06,840 --> 00:16:12,180 going to talk about, what you also have 298 00:16:12,180 --> 00:16:13,725 is prenylated proteins. 299 00:16:16,670 --> 00:16:21,270 And prenylated proteins are shown here. 300 00:16:21,270 --> 00:16:25,230 So quite frequently, you have small little g 301 00:16:25,230 --> 00:16:28,140 proteins, GTPases, you've seen these before. 302 00:16:28,140 --> 00:16:31,230 EFTU, EFGG, they're all over the place. 303 00:16:31,230 --> 00:16:32,760 There are hundreds of g proteins-- 304 00:16:32,760 --> 00:16:38,220 we talked about them in the recitation section on Rodnina 305 00:16:38,220 --> 00:16:39,390 that I gave you. 306 00:16:39,390 --> 00:16:43,080 Anyhow, a lot of those little g proteins go to the membrane 307 00:16:43,080 --> 00:16:44,760 and come away from the membrane. 308 00:16:44,760 --> 00:16:47,160 They do that by sticking on a tag. 309 00:16:47,160 --> 00:16:51,820 This tag can be geranylated or gerynalgeranylated-- 310 00:16:51,820 --> 00:16:53,850 it just the hydrophobic tag that allows 311 00:16:53,850 --> 00:16:57,720 things to interact with the membrane, 312 00:16:57,720 --> 00:17:00,960 increasing the effective molarity. 313 00:17:00,960 --> 00:17:04,890 Nature uses this trick over and over and over again. 314 00:17:04,890 --> 00:17:12,170 Another thing you can generate is natural products 315 00:17:12,170 --> 00:17:13,620 of medicinal interest. 316 00:17:18,750 --> 00:17:25,204 And I just show here taxol and artemisinin. 317 00:17:29,150 --> 00:17:35,900 So taxol is used in the treatment of breast cancer. 318 00:17:35,900 --> 00:17:39,300 Artemisinin, anybody heard of that? 319 00:17:39,300 --> 00:17:41,760 Yeah, so this has been the major target-- 320 00:17:41,760 --> 00:17:43,830 in fact, this pathway we're talking about today 321 00:17:43,830 --> 00:17:47,520 has been a major focus of many synthetic biologies, 322 00:17:47,520 --> 00:17:50,490 trying to make this mevalonic acid pathway 323 00:17:50,490 --> 00:17:56,100 so they can make potential drugs, but also jet fuels-- 324 00:17:56,100 --> 00:17:58,490 which, again, you want some kind of hydrocarbon. 325 00:17:58,490 --> 00:18:01,800 So this pathway has been studied a lot 326 00:18:01,800 --> 00:18:06,180 from a point of view of metabolic engineering. 327 00:18:06,180 --> 00:18:10,290 It's also involved in-- 328 00:18:10,290 --> 00:18:12,600 this is one of my favorite-- the perfume. 329 00:18:12,600 --> 00:18:18,120 You can tell from the way I smell the perfume industry. 330 00:18:18,120 --> 00:18:21,110 Any of you ever break a pine cone? 331 00:18:21,110 --> 00:18:22,500 A pine needle? 332 00:18:22,500 --> 00:18:24,670 Yeah, doesn't it smell great? 333 00:18:24,670 --> 00:18:25,772 No, you don't think so? 334 00:18:25,772 --> 00:18:26,730 I think it's wonderful. 335 00:18:26,730 --> 00:18:27,630 It's called pinene. 336 00:18:27,630 --> 00:18:28,560 Anyhow, it has-- 337 00:18:28,560 --> 00:18:30,580 I think terpenes wonderful smells, 338 00:18:30,580 --> 00:18:34,800 and it's the hallmark of the fragrance industry. 339 00:18:34,800 --> 00:18:35,510 Is that on here? 340 00:18:35,510 --> 00:18:36,720 Yeah, so menthol. 341 00:18:36,720 --> 00:18:41,460 Limonene is orange-- in fact, if you were here when Barry 342 00:18:41,460 --> 00:18:42,630 Sharpless used to teach-- 343 00:18:42,630 --> 00:18:44,572 I can't digress, because that's why I never 344 00:18:44,572 --> 00:18:45,780 get to the end of the course. 345 00:18:45,780 --> 00:18:49,470 But anyhow, Barry used to bring to organic class-- 346 00:18:49,470 --> 00:18:51,870 he had boxes of smells. 347 00:18:51,870 --> 00:18:53,820 And he used to pass around the smells, 348 00:18:53,820 --> 00:18:55,230 and they were all wonderful. 349 00:18:55,230 --> 00:18:59,280 And they were almost all terpenes. 350 00:18:59,280 --> 00:19:05,130 And we're going to be looking at things 351 00:19:05,130 --> 00:19:07,620 like dolichol-- we aren't going to be looking at it. 352 00:19:07,620 --> 00:19:10,200 We will see it a little bit. 353 00:19:10,200 --> 00:19:13,500 But what you can see here, Suzanne Walker 354 00:19:13,500 --> 00:19:15,900 is giving a talk here April 4th. 355 00:19:15,900 --> 00:19:17,160 And she works on-- 356 00:19:17,160 --> 00:19:20,340 one of the things she works on is peptidoglycan biosynthesis. 357 00:19:20,340 --> 00:19:24,990 And so sugars are carried around on these lipids. 358 00:19:24,990 --> 00:19:28,370 Some of them are C19 to C55. 359 00:19:28,370 --> 00:19:31,320 If you look at these, you can see these little units 360 00:19:31,320 --> 00:19:32,190 stuck together. 361 00:19:32,190 --> 00:19:35,940 Barbara Imperiali, in our department, the biology 362 00:19:35,940 --> 00:19:38,940 department, works on a asparagine-linked 363 00:19:38,940 --> 00:19:40,650 glycosylation. 364 00:19:40,650 --> 00:19:43,420 Again, the sugars are carried around 365 00:19:43,420 --> 00:19:45,230 by these kinds of terpenes. 366 00:19:45,230 --> 00:19:51,820 So plays a central role in putting sugars onto systems. 367 00:19:51,820 --> 00:19:56,360 And then what we're going to be focused on today-- 368 00:19:56,360 --> 00:19:59,380 and this is the focus in general-- 369 00:19:59,380 --> 00:20:02,605 is on cholesterol. 370 00:20:02,605 --> 00:20:03,910 Do I have that up there? 371 00:20:03,910 --> 00:20:05,350 I think so. 372 00:20:05,350 --> 00:20:07,730 So what we're going to-- do I have cholesterol up there? 373 00:20:07,730 --> 00:20:08,285 Yeah. 374 00:20:08,285 --> 00:20:08,910 OK, here it is. 375 00:20:08,910 --> 00:20:09,970 Cholesterol. 376 00:20:09,970 --> 00:20:12,110 That's what we're going to be focusing on. 377 00:20:12,110 --> 00:20:15,350 And that's not a C5, But a C30. 378 00:20:15,350 --> 00:20:21,290 So how do we get from these C5 units into the C30 units? 379 00:20:21,290 --> 00:20:26,240 So that's an introduction to the terpenome. 380 00:20:26,240 --> 00:20:27,720 They're everywhere. 381 00:20:27,720 --> 00:20:30,420 And so you can't become a biochemist 382 00:20:30,420 --> 00:20:34,710 without seeing carbon carbon bond formation by these C5 383 00:20:34,710 --> 00:20:37,740 units, in many, many kinds of reactions-- 384 00:20:37,740 --> 00:20:40,470 in both primary and secondary metabolism. 385 00:20:40,470 --> 00:20:42,860 They're very important. 386 00:20:42,860 --> 00:20:45,090 So what I want to do before we get 387 00:20:45,090 --> 00:20:48,600 into looking at one of the pathways 388 00:20:48,600 --> 00:20:54,390 that you can make the building blocks, IPP and DMAPP-- 389 00:20:57,350 --> 00:21:00,330 this is abbreviated DMAPP. 390 00:21:05,210 --> 00:21:08,210 One of the ways is through the mevalonic acid pathway, 391 00:21:08,210 --> 00:21:11,780 and here's a picture of a cell that I took from something 392 00:21:11,780 --> 00:21:12,860 off the web. 393 00:21:12,860 --> 00:21:16,250 But I want to introduce you to where we're going to be going, 394 00:21:16,250 --> 00:21:18,680 cholesterol biosynthesis. 395 00:21:18,680 --> 00:21:22,100 So where do we break down fatty acids? 396 00:21:25,100 --> 00:21:25,940 Does anybody know? 397 00:21:25,940 --> 00:21:28,550 You remember from your introductory course? 398 00:21:28,550 --> 00:21:31,760 I want to try to put this into the big picture on metabolism, 399 00:21:31,760 --> 00:21:35,270 so you're not-- we're just not pulling it out of the air. 400 00:21:35,270 --> 00:21:38,570 Anybody know where you break down fatty acids from the diet? 401 00:21:41,986 --> 00:21:44,069 AUDIENCE: You're asking in the cell, specifically? 402 00:21:44,069 --> 00:21:45,444 JOANNE STUBBE: There in the cell. 403 00:21:45,444 --> 00:21:46,490 Yeah, where in the cell? 404 00:21:46,490 --> 00:21:49,420 These are-- we're talking about eukaryotes now. 405 00:21:49,420 --> 00:21:50,760 Bacteria don't make cholesterol. 406 00:21:50,760 --> 00:21:51,260 Yeah. 407 00:21:51,260 --> 00:21:53,330 What? 408 00:21:53,330 --> 00:21:54,540 OK, you don't even know that. 409 00:21:54,540 --> 00:21:57,610 OK, so you should go back and read the chapter 410 00:21:57,610 --> 00:22:00,760 on fatty acid biosynthesis and degradation. 411 00:22:00,760 --> 00:22:02,920 That would be a good thing for you to do. 412 00:22:02,920 --> 00:22:06,520 Anyhow, fatty acids are broken down in the mitochondria. 413 00:22:06,520 --> 00:22:08,150 We'll see this in a second. 414 00:22:08,150 --> 00:22:10,510 So the mitochondria play a role. 415 00:22:10,510 --> 00:22:12,670 We're going to see today-- 416 00:22:12,670 --> 00:22:17,110 so here's the nucleus, here's the endoplasmic reticulum. 417 00:22:17,110 --> 00:22:20,830 The endoplasmic reticulum is the key sensor 418 00:22:20,830 --> 00:22:23,830 in cholesterol homeostasis. 419 00:22:23,830 --> 00:22:25,930 And so we're going to-- and we're 420 00:22:25,930 --> 00:22:29,410 going to see that it controls transcription factors. 421 00:22:29,410 --> 00:22:33,160 Transcription factors are stuck in a membrane in the ER. 422 00:22:33,160 --> 00:22:34,590 That's-- how weird is that? 423 00:22:34,590 --> 00:22:38,600 Because where do transcription factors need to go? 424 00:22:38,600 --> 00:22:40,582 They need to go to the nucleus. 425 00:22:40,582 --> 00:22:41,540 So how can you do that? 426 00:22:41,540 --> 00:22:43,457 How can you take something stuck in a membrane 427 00:22:43,457 --> 00:22:44,870 and get it to the nucleus? 428 00:22:44,870 --> 00:22:48,740 So they need to go through a golgi stack. 429 00:22:48,740 --> 00:22:51,650 They do some stuff we're going to learn about 430 00:22:51,650 --> 00:22:54,830 to eventually get into the nucleus, where they control 431 00:22:54,830 --> 00:22:59,030 not only the levels of cholesterol proteins, 432 00:22:59,030 --> 00:23:03,560 but also of metabolism of phospholipids, 433 00:23:03,560 --> 00:23:05,120 triacylglycerols. 434 00:23:05,120 --> 00:23:09,182 so this takes us into the big realm of all lipid metabolism, 435 00:23:09,182 --> 00:23:10,640 which most of the time people don't 436 00:23:10,640 --> 00:23:13,280 spend a lot of time talking about 437 00:23:13,280 --> 00:23:15,320 in an introductory course. 438 00:23:15,320 --> 00:23:17,840 And I mean, one of the interesting questions-- 439 00:23:17,840 --> 00:23:21,920 we're going to see the key rate limiting step 440 00:23:21,920 --> 00:23:24,910 in cholesterol homeostasis. 441 00:23:24,910 --> 00:23:29,120 The protein is bound to the ER membrane, 442 00:23:29,120 --> 00:23:30,800 and a lot of the proteins involved 443 00:23:30,800 --> 00:23:35,150 in cholesterol biosynthesis are in the ER membrane. 444 00:23:35,150 --> 00:23:40,230 50% of all the cholesterol ends up in the plasma membrane. 445 00:23:40,230 --> 00:23:41,600 how does it get there? 446 00:23:41,600 --> 00:23:43,250 Does it just go through solution? 447 00:23:43,250 --> 00:23:46,590 You need to think about the properties of cholesterol. 448 00:23:46,590 --> 00:23:48,770 So when you get confused about where we're going, 449 00:23:48,770 --> 00:23:50,270 come back to the picture. 450 00:23:50,270 --> 00:23:54,260 And I'm going to show you one other big picture, which 451 00:23:54,260 --> 00:23:55,920 we use when I teach-- 452 00:23:55,920 --> 00:23:58,760 when I've taught 5.07 with Essigmann-- again, 453 00:23:58,760 --> 00:24:00,470 this is the picture we can back to over 454 00:24:00,470 --> 00:24:02,120 and over and over again. 455 00:24:02,120 --> 00:24:04,700 Because everything is interconnected. 456 00:24:04,700 --> 00:24:08,540 So we're going to be talking about cholesterol biosynthesis. 457 00:24:08,540 --> 00:24:12,300 We're going to see a key player is acetyl-CoA. 458 00:24:12,300 --> 00:24:13,430 Where have you seen that? 459 00:24:13,430 --> 00:24:17,130 You've learned a lot about acetyl-CoA in biosynthesis, 460 00:24:17,130 --> 00:24:20,460 and use in biosynthesis and polyketide. 461 00:24:20,460 --> 00:24:26,690 Synthases, you talked about biosynthesis of fatty acids. 462 00:24:26,690 --> 00:24:30,470 So fatty acids are biosynthesized in the cytosol. 463 00:24:33,500 --> 00:24:37,040 But I just told you fatty acids are 464 00:24:37,040 --> 00:24:40,970 degraded in the mitochondria. 465 00:24:40,970 --> 00:24:42,350 What are they degraded to? 466 00:24:42,350 --> 00:24:45,170 Degraded to acetyl-CoA. 467 00:24:45,170 --> 00:24:50,450 Can acetyl-CoA get from the mitochondria to the cytosol? 468 00:24:58,820 --> 00:25:02,030 Nobody knows? 469 00:25:02,030 --> 00:25:04,910 Let's get some energy, you guys. 470 00:25:04,910 --> 00:25:07,514 What do we know about acetyl-CoA? 471 00:25:07,514 --> 00:25:09,958 AUDIENCE: [INAUDIBLE] 472 00:25:09,958 --> 00:25:11,000 JOANNE STUBBE: It's what? 473 00:25:11,000 --> 00:25:13,295 AUDIENCE: A transport system [INAUDIBLE]---- 474 00:25:13,295 --> 00:25:15,128 JOANNE STUBBE: So you think is the transport 475 00:25:15,128 --> 00:25:17,850 system that takes it from the mitochondria to the cytosol. 476 00:25:17,850 --> 00:25:19,910 So that's wrong. 477 00:25:19,910 --> 00:25:21,890 And in fact, this is again another thing 478 00:25:21,890 --> 00:25:24,380 that I think maybe isn't emphasized enough 479 00:25:24,380 --> 00:25:25,850 in an introductory course. 480 00:25:25,850 --> 00:25:28,460 A lot of these things cannot transfer across these 481 00:25:28,460 --> 00:25:29,640 membranes. 482 00:25:29,640 --> 00:25:31,880 So-- and this may or may not be logical to you, 483 00:25:31,880 --> 00:25:33,020 when you take this and you're saying, 484 00:25:33,020 --> 00:25:34,437 oh my god, this is so complicated. 485 00:25:34,437 --> 00:25:36,110 It's really not that complicated when 486 00:25:36,110 --> 00:25:40,600 you put all primary metabolism into the big picture. 487 00:25:40,600 --> 00:25:44,180 Acetyl-CoA goes into the TCA cycle, 488 00:25:44,180 --> 00:25:46,790 and it condenses with oxaloacetic acid 489 00:25:46,790 --> 00:25:47,960 to form citric acid. 490 00:25:47,960 --> 00:25:51,440 We're going to see citric acid again with iron homeostasis. 491 00:25:51,440 --> 00:25:55,190 Anyhow, it's citrate that is able to go 492 00:25:55,190 --> 00:25:57,050 across the mitochondrial membrane, 493 00:25:57,050 --> 00:26:00,310 as is malate, as is pyruvate. 494 00:26:00,310 --> 00:26:03,260 Acetyl-CoA is not able to do that. 495 00:26:03,260 --> 00:26:05,420 And so to get acetyl-CoA, then you 496 00:26:05,420 --> 00:26:08,228 have to enzymatically break down citrate. 497 00:26:08,228 --> 00:26:09,770 So if you don't know what citrate is, 498 00:26:09,770 --> 00:26:11,030 it's a central metabolite. 499 00:26:11,030 --> 00:26:12,770 You're going to see it again and again. 500 00:26:12,770 --> 00:26:14,150 Pull it up, Google it. 501 00:26:14,150 --> 00:26:16,280 Put it in your brain. 502 00:26:16,280 --> 00:26:18,890 You form acetyl-CoA. 503 00:26:18,890 --> 00:26:22,260 An acetyl-CoA, as you learned in the first part of course, 504 00:26:22,260 --> 00:26:24,260 can form fatty acids. 505 00:26:24,260 --> 00:26:26,180 Where do fatty acids go? 506 00:26:26,180 --> 00:26:29,450 They can attach to glycerol. 507 00:26:29,450 --> 00:26:31,230 And where does glycerol come from? 508 00:26:31,230 --> 00:26:32,660 It comes from breakdown of sugars 509 00:26:32,660 --> 00:26:35,240 through the glycolosis pathway. 510 00:26:35,240 --> 00:26:39,017 And they come together to form phospholipids, which 511 00:26:39,017 --> 00:26:40,100 make up all our membranes. 512 00:26:40,100 --> 00:26:42,020 Pretty important. 513 00:26:42,020 --> 00:26:44,030 What else can fatty acids do? 514 00:26:44,030 --> 00:26:47,970 They can interact with glycerol without a phosphate, 515 00:26:47,970 --> 00:26:50,840 to triacylglycerols. 516 00:26:50,840 --> 00:26:54,020 Triacyl-- esterified triacylgycerols. 517 00:26:54,020 --> 00:26:57,400 Does everybody know what glycerol is? 518 00:26:57,400 --> 00:26:59,210 Everybody know? 519 00:26:59,210 --> 00:27:02,490 OK, so-- and this is another thing we're going to find. 520 00:27:02,490 --> 00:27:04,820 We have huge amounts of phospholipids 521 00:27:04,820 --> 00:27:07,970 and triacylglycerols in our diet. 522 00:27:07,970 --> 00:27:10,490 So we have to deal with those things. 523 00:27:10,490 --> 00:27:14,420 But acetyl-CoA, we'll also see, is the building block 524 00:27:14,420 --> 00:27:17,210 to form mevalonic acid through a pathway we're 525 00:27:17,210 --> 00:27:18,890 going to describe now. 526 00:27:18,890 --> 00:27:23,040 So mevalonic acid is a key player, 527 00:27:23,040 --> 00:27:25,100 and its formation is rate limiting 528 00:27:25,100 --> 00:27:27,620 in cholesterol biosynthesis. 529 00:27:27,620 --> 00:27:29,990 The enzyme that makes mevaloinc acid 530 00:27:29,990 --> 00:27:33,920 is located in this little messy thing here, and that's the ER. 531 00:27:33,920 --> 00:27:36,020 So it's bound to the ER. 532 00:27:36,020 --> 00:27:38,070 It makes cholesterol. 533 00:27:38,070 --> 00:27:42,530 And then ultimately, how does cholesterol move-- 534 00:27:42,530 --> 00:27:45,200 and a lot of the precursors to cholesterol stay 535 00:27:45,200 --> 00:27:47,720 solubilized, and then get distributed 536 00:27:47,720 --> 00:27:49,520 to all the membranes. 537 00:27:49,520 --> 00:27:51,830 50% of the cholesterol, for example, 538 00:27:51,830 --> 00:27:54,800 is found in the plasma membrane. 539 00:27:54,800 --> 00:27:56,200 So that's the big picture. 540 00:27:56,200 --> 00:27:59,690 And so when you get confused about where we're going, 541 00:27:59,690 --> 00:28:02,570 you need to go back and see how central a player 542 00:28:02,570 --> 00:28:06,260 acetyl-CoA is to everything. 543 00:28:06,260 --> 00:28:09,350 And so its regulation, you're going to see, 544 00:28:09,350 --> 00:28:11,930 is governed by the same transcription 545 00:28:11,930 --> 00:28:15,562 factors that regulate cholesterol homeostasis. 546 00:28:15,562 --> 00:28:16,770 Because they were all linked. 547 00:28:19,310 --> 00:28:20,513 So where are we going? 548 00:28:20,513 --> 00:28:22,430 Let's see if I can remember where we're going. 549 00:28:29,570 --> 00:28:30,650 So where are we going? 550 00:28:35,330 --> 00:28:40,580 So I'm giving you an overview of where we're going. 551 00:28:40,580 --> 00:28:46,010 And I'm only-- this is a long pathway to get to lanosterol. 552 00:28:46,010 --> 00:28:48,380 I'm not going to look at all the steps in the pathway. 553 00:28:48,380 --> 00:28:52,280 I'm just going to tell you how we use these C5 554 00:28:52,280 --> 00:28:54,440 units to generate terpenes. 555 00:28:54,440 --> 00:28:56,840 So we've got to get to the C5 units over here. 556 00:28:56,840 --> 00:29:00,350 We've got to get to these two intermediates. 557 00:29:00,350 --> 00:29:05,060 And then we're going to use them to get eventually to a C30. 558 00:29:05,060 --> 00:29:06,710 And we'll see the same chemistry, 559 00:29:06,710 --> 00:29:10,070 once we know a few rules, just like with aldol reactions 560 00:29:10,070 --> 00:29:14,540 and claison reactions, are used over and over and over again. 561 00:29:14,540 --> 00:29:15,800 There are a few basic rules. 562 00:29:15,800 --> 00:29:17,840 Every protein is different, but I'm 563 00:29:17,840 --> 00:29:20,240 going to make sweeping generalizations, which 564 00:29:20,240 --> 00:29:22,390 is a good place to start. 565 00:29:22,390 --> 00:29:29,510 So we have to use three molecules of acetyl-CoA. 566 00:29:29,510 --> 00:29:31,250 So you're all-- you all should be 567 00:29:31,250 --> 00:29:34,520 very familiar with acetyl-CoA. 568 00:29:34,520 --> 00:29:41,160 And we're after trying to form C5. 569 00:29:41,160 --> 00:29:44,700 So three of these gives us C6, so we 570 00:29:44,700 --> 00:29:46,730 have to get rid of a carbon. 571 00:29:46,730 --> 00:29:51,600 So from this we need to lose whatever the pathway is. 572 00:29:51,600 --> 00:29:55,410 It turns out we lose one carbon as CO2. 573 00:29:55,410 --> 00:29:56,650 And this whole process-- 574 00:29:56,650 --> 00:29:59,510 so this can be multiple steps-- 575 00:29:59,510 --> 00:30:02,610 is called initiation. 576 00:30:02,610 --> 00:30:09,120 And it forms IPP, which can isomerize to form DMAPP. 577 00:30:11,940 --> 00:30:15,390 And then, again, this is our C5 unit that we're after. 578 00:30:15,390 --> 00:30:19,020 So this is C5. 579 00:30:19,020 --> 00:30:24,840 And we've lost one carbon as CO2. 580 00:30:24,840 --> 00:30:29,040 So then we're going to have what I'm going to call elongation. 581 00:30:32,940 --> 00:30:38,050 And what we're going to see is that to get to lanosterol, 582 00:30:38,050 --> 00:30:41,200 all we need to have a C30. 583 00:30:41,200 --> 00:30:42,740 So we need six of these guys. 584 00:30:42,740 --> 00:30:48,680 So we have six C5 to form a C30. 585 00:30:53,540 --> 00:30:54,630 Which is lanosterol. 586 00:30:54,630 --> 00:30:57,770 This is a precursor to steroids. 587 00:30:57,770 --> 00:31:03,980 [INAUDIBLE] talk about cholesterol-- uh oh. 588 00:31:03,980 --> 00:31:07,520 I knew that that was going to happen. 589 00:31:07,520 --> 00:31:08,180 I jump around. 590 00:31:08,180 --> 00:31:10,020 Usually this falls off, so you'll 591 00:31:10,020 --> 00:31:12,820 have to get used to this. 592 00:31:12,820 --> 00:31:13,660 It's a good thing-- 593 00:31:13,660 --> 00:31:18,374 I spent all morning trying to figure out where the-- 594 00:31:18,374 --> 00:31:20,650 where this cord came, because I knew 595 00:31:20,650 --> 00:31:23,310 my cord wasn't going to fit, and the cord was going to be shot, 596 00:31:23,310 --> 00:31:25,810 and then I was thinking, how am I going to run from one door 597 00:31:25,810 --> 00:31:27,070 to the next? 598 00:31:27,070 --> 00:31:31,210 I need to get this back on me. 599 00:31:31,210 --> 00:31:32,140 Can you hear me? 600 00:31:32,140 --> 00:31:32,640 OK. 601 00:31:36,990 --> 00:31:37,990 Let me get back on gear. 602 00:31:37,990 --> 00:31:40,540 So C30-- but then what's going to happen-- 603 00:31:40,540 --> 00:31:44,650 so we get to lanosterol, and then from lanosterol, 604 00:31:44,650 --> 00:31:49,930 and going to lanosterol here, we're going to have to do, 605 00:31:49,930 --> 00:31:52,690 like, I think, the most amazing chemistry in the whole world. 606 00:31:52,690 --> 00:31:59,650 We're going to have to do an oxidation and a cyclization. 607 00:31:59,650 --> 00:32:01,330 So this is going to be a terpene. 608 00:32:01,330 --> 00:32:03,435 So we're going to put these things together 609 00:32:03,435 --> 00:32:06,220 to form a C30, a linear C30, and then 610 00:32:06,220 --> 00:32:10,120 they have to come together to form this guy. 611 00:32:10,120 --> 00:32:12,640 So we're going to talk about this reaction, 612 00:32:12,640 --> 00:32:14,650 because it's such a cool reaction. 613 00:32:14,650 --> 00:32:17,800 Anyhow, we're going to talk about how this linear molecule 614 00:32:17,800 --> 00:32:18,790 gets to this. 615 00:32:18,790 --> 00:32:22,840 That's-- that is the coolest reaction, in my opinion, 616 00:32:22,840 --> 00:32:23,650 in biology. 617 00:32:23,650 --> 00:32:26,500 I remember when I first heard about this when I 618 00:32:26,500 --> 00:32:29,500 was in graduate school in 1968. 619 00:32:29,500 --> 00:32:30,640 A long time ago. 620 00:32:30,640 --> 00:32:32,200 That's what made me decide I didn't 621 00:32:32,200 --> 00:32:34,270 want to be an organic chemist. 622 00:32:34,270 --> 00:32:37,480 I said, how amazing is this? 623 00:32:37,480 --> 00:32:40,600 That you can do one step and you can 624 00:32:40,600 --> 00:32:45,580 put in all of these asymmetric centers and 100% yield. 625 00:32:45,580 --> 00:32:46,360 So that was it. 626 00:32:46,360 --> 00:32:48,520 That was a turning point in my life. 627 00:32:48,520 --> 00:32:51,490 Anyhow, hopefully it'll be a turning point in your life too. 628 00:32:51,490 --> 00:32:53,930 So we have C30. 629 00:32:53,930 --> 00:32:56,150 And then we're not there yet. 630 00:32:56,150 --> 00:32:58,420 So this is going to be, for us, the-- 631 00:32:58,420 --> 00:33:02,500 after the elongation, these cyclizations and oxidation 632 00:33:02,500 --> 00:33:07,970 is going to be the termination to get to this ring structure. 633 00:33:07,970 --> 00:33:12,490 But then to get to cholesterol, we have 19 more steps. 634 00:33:15,610 --> 00:33:18,910 So this is really complicated pathway. 635 00:33:18,910 --> 00:33:21,760 And I'll tell you what had the chemistry actually is not 636 00:33:21,760 --> 00:33:24,370 so hard to understand, but the details are really 637 00:33:24,370 --> 00:33:25,720 still not understood. 638 00:33:25,720 --> 00:33:30,710 Because all of the proteins are membrane bound. 639 00:33:30,710 --> 00:33:34,360 So what I want to do now is come back over here. 640 00:33:34,360 --> 00:33:39,080 And we're going to talk about initiation, elongation, 641 00:33:39,080 --> 00:33:40,450 and the termination steps. 642 00:33:40,450 --> 00:33:43,570 And I'm going to focus on a few of the reactions 643 00:33:43,570 --> 00:33:48,190 that I think are important, and a lot of the details 644 00:33:48,190 --> 00:33:50,120 are written down-- 645 00:33:50,120 --> 00:33:54,100 are written down on the PowerPoint. 646 00:33:54,100 --> 00:33:57,130 So let's look at the first few steps. 647 00:33:57,130 --> 00:33:59,455 And so let's start the pathway. 648 00:34:02,230 --> 00:34:08,484 And the first molecule we're dealing with is acetyl-CoA. 649 00:34:11,040 --> 00:34:13,860 And what is special about acetyl-CoA. 650 00:34:13,860 --> 00:34:17,370 Why is nature-- you've just had a whole bunch of units 651 00:34:17,370 --> 00:34:23,880 on acetyl-CoA-- why does nature use thioesters? 652 00:34:23,880 --> 00:34:25,560 What are the two key things you need 653 00:34:25,560 --> 00:34:28,230 to think about in terms of its reactivity? 654 00:34:28,230 --> 00:34:30,280 You guys should be experts on this now. 655 00:34:30,280 --> 00:34:30,780 Yeah? 656 00:34:33,842 --> 00:34:35,550 AUDIENCE: There's a low pKa [INAUDIBLE].. 657 00:34:35,550 --> 00:34:37,699 JOANNE STUBBE: Right, so you have a reduced pKa, 658 00:34:37,699 --> 00:34:40,850 is reduced from, say, 22 to 18. 659 00:34:40,850 --> 00:34:44,239 So this is the alpha hydrogen acidity. 660 00:34:44,239 --> 00:34:48,790 And what else does a thioester do? 661 00:34:48,790 --> 00:34:52,550 What is the other reactive part of CoA? 662 00:34:52,550 --> 00:34:54,929 This should be like the back of your hand. 663 00:34:54,929 --> 00:34:57,450 I mean, this is part-- this is central to everything 664 00:34:57,450 --> 00:34:58,300 in biochemistry. 665 00:34:58,300 --> 00:34:58,800 Yeah. 666 00:34:58,800 --> 00:35:00,230 AUDIENCE: [INAUDIBLE] 667 00:35:00,230 --> 00:35:00,670 JOANNE STUBBE: Pardon me? 668 00:35:00,670 --> 00:35:01,837 AUDIENCE: The leaving group. 669 00:35:01,837 --> 00:35:04,800 JOANNE STUBBE: The leaving group. 670 00:35:04,800 --> 00:35:06,480 You are going to have a leaving group. 671 00:35:06,480 --> 00:35:08,610 But that's-- and that is important, 672 00:35:08,610 --> 00:35:11,250 but that's not the key important thing. 673 00:35:11,250 --> 00:35:13,015 That is a part of the game. 674 00:35:13,015 --> 00:35:14,640 It can drive the reaction to the right, 675 00:35:14,640 --> 00:35:17,880 if you look at the free energy of hydrolysis. 676 00:35:17,880 --> 00:35:22,740 What else is activated when you have a sulfur ester as opposed 677 00:35:22,740 --> 00:35:24,220 to an oxygen ester? 678 00:35:24,220 --> 00:35:25,400 AUDIENCE: Carbonyl. 679 00:35:25,400 --> 00:35:27,930 JOANNE STUBBE: The carbonyl, because of the decreased 680 00:35:27,930 --> 00:35:29,910 resonance stabilization. 681 00:35:29,910 --> 00:35:33,450 So what you've done then is you have activation 682 00:35:33,450 --> 00:35:35,880 for nucleophilic attack. 683 00:35:35,880 --> 00:35:38,010 And you see this-- nature uses this 684 00:35:38,010 --> 00:35:41,190 in cholesterol homeostasis as well, 685 00:35:41,190 --> 00:35:42,900 over and over and over again. 686 00:35:42,900 --> 00:35:45,360 So in the first step, and I'm not going to draw out 687 00:35:45,360 --> 00:35:50,550 the details, what you can see here is that you're taking two 688 00:35:50,550 --> 00:35:54,600 molecules of acetyl-CoA and you're forming 689 00:35:54,600 --> 00:35:56,010 acetoacetyl-CoA-- 690 00:35:56,010 --> 00:35:57,690 that should be good practice for you 691 00:35:57,690 --> 00:36:02,300 for thinking about the exam on Wednesday. 692 00:36:05,520 --> 00:36:08,480 And this is an example of a claison reaction, 693 00:36:08,480 --> 00:36:13,460 one of the three types of mechanisms, 694 00:36:13,460 --> 00:36:16,000 to form carbon carbon bonds. 695 00:36:16,000 --> 00:36:20,570 The next step, we need three acetyl-CoA's 696 00:36:20,570 --> 00:36:25,010 to get eventually to isopentenyl pyrophosphate and dimethylallyl 697 00:36:25,010 --> 00:36:25,940 pyrophosphate. 698 00:36:25,940 --> 00:36:29,770 So we're going to use another molecule on acetyl-CoA 699 00:36:29,770 --> 00:36:34,790 to form hydroxymethylglutaryl-CoA. 700 00:36:34,790 --> 00:36:37,430 So we need to add another one of these guys. 701 00:36:40,010 --> 00:36:42,030 And this is HMG-CoA synthase. 702 00:36:45,850 --> 00:36:48,870 So before I go there, let's go through what we know. 703 00:36:48,870 --> 00:36:53,740 So this is-- so we're starting here. 704 00:36:53,740 --> 00:36:57,640 So here-- acetyl-CoA plays a central role. 705 00:36:57,640 --> 00:36:58,810 Why thioesters? 706 00:36:58,810 --> 00:37:01,080 It's important in claison reactions. 707 00:37:01,080 --> 00:37:04,120 Here's the example of a claison reaction involving 708 00:37:04,120 --> 00:37:08,780 a carbanion intermediate that you guys should all be experts 709 00:37:08,780 --> 00:37:10,870 at at this stage. 710 00:37:10,870 --> 00:37:11,900 What about this step? 711 00:37:11,900 --> 00:37:12,700 The next step? 712 00:37:12,700 --> 00:37:16,690 Formation of hydroxymethylglutaryl-CoA? 713 00:37:16,690 --> 00:37:22,180 So here it turns out that this enzyme uses covalent catalysis. 714 00:37:22,180 --> 00:37:24,478 Frequently enzymes-- you've already seen this as well, 715 00:37:24,478 --> 00:37:26,770 we're going to see this again and again over the course 716 00:37:26,770 --> 00:37:28,270 of the rest of the semester. 717 00:37:28,270 --> 00:37:30,730 One of the major mechanisms of rate acceleration 718 00:37:30,730 --> 00:37:32,460 is covalent catalysis. 719 00:37:32,460 --> 00:37:36,340 Here the thioester-- the CoA ester has been removed 720 00:37:36,340 --> 00:37:40,090 and it's attached to a cysteine in the active site 721 00:37:40,090 --> 00:37:41,140 of the enzyme. 722 00:37:41,140 --> 00:37:44,170 And then this can react with, in this case, 723 00:37:44,170 --> 00:37:48,080 a ketone like molecule in an aldol reaction. 724 00:37:48,080 --> 00:37:50,830 So here's the second example. 725 00:37:50,830 --> 00:37:52,430 We take acetoacetyl-CoA. 726 00:37:58,000 --> 00:37:59,680 We add another CoA. 727 00:37:59,680 --> 00:38:04,160 And what we form, then, is hydroxymethylglutaryl-CoA. 728 00:38:06,850 --> 00:38:10,390 And what we're going to see is, during this reaction, 729 00:38:10,390 --> 00:38:14,620 we also have to do a hydrolosis reaction. 730 00:38:14,620 --> 00:38:18,160 Because we start out with acetyl-CoA and we only add-- 731 00:38:18,160 --> 00:38:21,550 end up with a single thioester. 732 00:38:24,230 --> 00:38:31,050 And this reaction forms hydroxymethylglutaryl-CoA. 733 00:38:35,310 --> 00:38:37,980 So we've lost-- in this reaction over here, 734 00:38:37,980 --> 00:38:39,360 you can see where this is lost. 735 00:38:39,360 --> 00:38:43,950 So you have an acetyl-CoA to form the thioester 736 00:38:43,950 --> 00:38:45,550 in the active site. 737 00:38:45,550 --> 00:38:47,550 You you've lost a CoA. 738 00:38:47,550 --> 00:38:49,330 Is everybody with me? 739 00:38:49,330 --> 00:38:52,560 So you're using the third molecule of acetyl-CoA. 740 00:38:52,560 --> 00:38:54,900 You've already lost the CoA. 741 00:38:54,900 --> 00:38:57,570 And then what you end up with in the end 742 00:38:57,570 --> 00:39:00,490 is you have to hydrolize this off. 743 00:39:00,490 --> 00:39:02,760 So if you didn't realize it went through a covalent 744 00:39:02,760 --> 00:39:04,470 intermediate, it would be like you just 745 00:39:04,470 --> 00:39:06,270 lost a CoA, which you did. 746 00:39:06,270 --> 00:39:09,420 But you lost it in this step, because you 747 00:39:09,420 --> 00:39:11,460 went through a covalent intermediate. 748 00:39:11,460 --> 00:39:13,890 And you're not responsible for the details. 749 00:39:13,890 --> 00:39:17,460 Many, many enzymes that use acetyl-CoA go through covalent 750 00:39:17,460 --> 00:39:19,600 intermediates just like this. 751 00:39:19,600 --> 00:39:23,170 But you have to study each one to figure out why they do that. 752 00:39:23,170 --> 00:39:24,150 Why do they do that? 753 00:39:24,150 --> 00:39:27,750 Because covalent catalysis gives us rate accelerations of 10 754 00:39:27,750 --> 00:39:29,710 to the 4th, 10 to the 5th. 755 00:39:29,710 --> 00:39:31,950 So nature has used that as a repertoire 756 00:39:31,950 --> 00:39:37,440 of defining how to catalyze reactions at amazing rates. 757 00:39:37,440 --> 00:39:39,930 So now we're at this stage. 758 00:39:39,930 --> 00:39:42,060 And the next step in this pathway-- 759 00:39:42,060 --> 00:39:46,020 so we're still trying to get to the C5 over here. 760 00:39:50,900 --> 00:39:53,510 And to get to this now, we're going 761 00:39:53,510 --> 00:39:56,370 to have to do a reduction. 762 00:39:56,370 --> 00:39:58,640 And we're trying to get to-- 763 00:39:58,640 --> 00:40:00,860 so we did this reaction here. 764 00:40:03,900 --> 00:40:05,410 I will fix my thing for next-- 765 00:40:05,410 --> 00:40:05,980 OK. 766 00:40:05,980 --> 00:40:08,710 So now what we're doing is we're going 767 00:40:08,710 --> 00:40:12,970 from hydroxymethylglutaryl-CoA to mevalonic acid. 768 00:40:12,970 --> 00:40:15,620 And this is wrong. 769 00:40:15,620 --> 00:40:17,990 They should all be NADPHs. 770 00:40:17,990 --> 00:40:20,720 When you're doing biosynthesis, what do you use? 771 00:40:20,720 --> 00:40:23,000 You don't use any NADH. 772 00:40:23,000 --> 00:40:27,980 You use NADPH in almost all biosynthetic pathways. 773 00:40:27,980 --> 00:40:29,240 So what happens? 774 00:40:29,240 --> 00:40:34,150 You're reducing, basically, the thioester down to-- 775 00:40:34,150 --> 00:40:37,910 a thioester down to an alcohol. 776 00:40:37,910 --> 00:40:41,030 Everybody should know at this stage how this kind of reaction 777 00:40:41,030 --> 00:40:42,620 goes. 778 00:40:42,620 --> 00:40:48,530 Everybody, this is one of the two major redox co-factors 779 00:40:48,530 --> 00:40:49,340 and all of biology. 780 00:40:53,910 --> 00:40:58,610 Can somebody tell me how this redox reaction goes? 781 00:40:58,610 --> 00:41:01,550 This is one of the vitamins on your bottle, niacin, 782 00:41:01,550 --> 00:41:05,840 which gets metabolized into NAD, NADP. 783 00:41:05,840 --> 00:41:09,230 How does that do a reduction? 784 00:41:09,230 --> 00:41:10,130 Can somebody tell me? 785 00:41:16,696 --> 00:41:17,640 Yeah. 786 00:41:17,640 --> 00:41:20,595 AUDIENCE: Form an aromatic ring by eliminating the hydride-- 787 00:41:20,595 --> 00:41:21,810 so the hydride attacks the-- 788 00:41:21,810 --> 00:41:23,268 JOANNE STUBBE: Right, so that's it. 789 00:41:23,268 --> 00:41:25,662 And where does the hydride attack? 790 00:41:25,662 --> 00:41:28,067 AUDIENCE: The carbonyl. 791 00:41:28,067 --> 00:41:29,775 JOANNE STUBBE: What part of the carbonyl? 792 00:41:29,775 --> 00:41:30,770 AUDIENCE: The carbon. 793 00:41:30,770 --> 00:41:31,770 JOANNE STUBBE: Yeah, OK. 794 00:41:31,770 --> 00:41:34,770 So this is something that I fight with kids all the time 795 00:41:34,770 --> 00:41:35,860 in 5.07. 796 00:41:35,860 --> 00:41:38,850 It doesn't attack the oxygen. It attacks the carbonyl 797 00:41:38,850 --> 00:41:42,840 because it's polarized delta plus delta minus. 798 00:41:42,840 --> 00:41:46,920 So you have-- this, again, of all the vitamins 799 00:41:46,920 --> 00:41:49,950 on your vitamin bottle, this is the simplest one. 800 00:41:49,950 --> 00:41:53,640 So hydrogen moves with a pair of electrons 801 00:41:53,640 --> 00:41:58,090 that's called the hydride, to do this reduction. 802 00:41:58,090 --> 00:42:03,300 And over here, I think I have the details written out. 803 00:42:03,300 --> 00:42:07,290 So I'm not going to write this out in more detail. 804 00:42:07,290 --> 00:42:11,070 But you generate this intermediate-- 805 00:42:11,070 --> 00:42:13,780 this intermediate-- 806 00:42:13,780 --> 00:42:17,790 this intermediate may-- or the oxygen may or may not 807 00:42:17,790 --> 00:42:18,570 be protonated. 808 00:42:18,570 --> 00:42:22,440 You need to look in the active site of the enzyme. 809 00:42:22,440 --> 00:42:25,590 But then what happens to this intermediate? 810 00:42:25,590 --> 00:42:28,320 This intermediate-- so tetrahedral intermediate's 811 00:42:28,320 --> 00:42:29,820 not very stable. 812 00:42:29,820 --> 00:42:33,750 It can break down to liberate CoA. 813 00:42:33,750 --> 00:42:34,990 And what are you left with? 814 00:42:34,990 --> 00:42:37,630 You're left with an aldehyde. 815 00:42:37,630 --> 00:42:39,520 So that's one reduction. 816 00:42:39,520 --> 00:42:41,410 And where do we want to go? 817 00:42:41,410 --> 00:42:43,270 Where we want to go-- 818 00:42:43,270 --> 00:42:45,400 and so I'm not going to draw the whole thing out, 819 00:42:45,400 --> 00:42:47,440 but I'll draw a part of this out-- 820 00:42:47,440 --> 00:42:53,080 so this gives us, then, through a tetrahedral intermediate, 821 00:42:53,080 --> 00:42:56,040 an aldehyde. 822 00:42:56,040 --> 00:43:00,630 And then what can happen to the aldehyde? 823 00:43:00,630 --> 00:43:03,270 We use another molecule of NADPH? 824 00:43:03,270 --> 00:43:04,530 And what happens with that? 825 00:43:04,530 --> 00:43:05,220 The same thing. 826 00:43:05,220 --> 00:43:08,250 You now do a hydride transfer. 827 00:43:08,250 --> 00:43:17,260 And so we need another molecule of NADPH to form the alcohol. 828 00:43:17,260 --> 00:43:20,650 So this is a mevalonic acid. 829 00:43:20,650 --> 00:43:24,160 So this is written out in more detail 830 00:43:24,160 --> 00:43:28,240 here, for those of you who have trouble trouble thinking 831 00:43:28,240 --> 00:43:28,750 about this. 832 00:43:28,750 --> 00:43:32,730 But of all the factors that nature 833 00:43:32,730 --> 00:43:36,390 has evolved to help us expand the repertoire of reactions 834 00:43:36,390 --> 00:43:38,040 in biology, NADPH-- 835 00:43:38,040 --> 00:43:41,540 NADH is the simplest. 836 00:43:41,540 --> 00:43:43,920 Hydride-- it's always hydride. 837 00:43:43,920 --> 00:43:46,050 Flavins, much more complicated. 838 00:43:46,050 --> 00:43:48,480 We'll see some of those-- the chemistry is much more 839 00:43:48,480 --> 00:43:49,080 complicated. 840 00:43:49,080 --> 00:43:53,450 This is really straightforward. 841 00:43:53,450 --> 00:43:55,700 So what do we know about this? 842 00:43:55,700 --> 00:43:58,030 Why are people interested in this? 843 00:43:58,030 --> 00:44:07,880 And this enzyme is called HMG-CoA reductase. 844 00:44:07,880 --> 00:44:11,630 And in your handouts, it's abbreviated-- 845 00:44:11,630 --> 00:44:14,320 I think these things are terrible. 846 00:44:14,320 --> 00:44:17,240 I will give you a list with all the acronyms on them. 847 00:44:17,240 --> 00:44:19,040 I can't remember the acronyms. 848 00:44:19,040 --> 00:44:20,450 And people change them. 849 00:44:20,450 --> 00:44:25,590 And people name things-- enzyme names are extremely difficult. 850 00:44:25,590 --> 00:44:29,270 The older they are, the worse the issue is. 851 00:44:29,270 --> 00:44:34,310 Because do you know what NAD used to be called? 852 00:44:34,310 --> 00:44:36,010 Any of you have a memory of that? 853 00:44:36,010 --> 00:44:38,083 Any of you read the old literature? 854 00:44:41,870 --> 00:44:44,310 It used to be called DPN-- 855 00:44:44,310 --> 00:44:47,780 dipyridine nucleotide. 856 00:44:47,780 --> 00:44:52,670 So this is pyridine, and that's where they got the name from. 857 00:44:52,670 --> 00:44:55,040 And I used to teach with somebody, [INAUDIBLE] 858 00:44:55,040 --> 00:44:58,010 a long time ago, and everything was DPN. 859 00:44:58,010 --> 00:44:59,780 So anyhow, if you've read the literature, 860 00:44:59,780 --> 00:45:02,030 nothing will be called in NAD, NADH. 861 00:45:02,030 --> 00:45:04,190 And in fact, a lot of the seminal experiments 862 00:45:04,190 --> 00:45:08,540 that elucidate the pathways came out of the old literature. 863 00:45:08,540 --> 00:45:10,760 So why is this protein interesting? 864 00:45:10,760 --> 00:45:13,400 So we're going to spend a little bit of time on this protein. 865 00:45:13,400 --> 00:45:18,170 People have spent a huge amount of time looking 866 00:45:18,170 --> 00:45:19,820 at this protein in detail. 867 00:45:19,820 --> 00:45:22,286 Does anybody know why? 868 00:45:22,286 --> 00:45:24,745 AUDIENCE: [INAUDIBLE] people target it-- like statins 869 00:45:24,745 --> 00:45:26,240 target it, or cholesterol. 870 00:45:26,240 --> 00:45:28,790 JOANNE STUBBE: So the key thing in this system 871 00:45:28,790 --> 00:45:35,600 is it's the rate limiting step in cholesterol biosynthesis. 872 00:45:38,330 --> 00:45:45,290 And it's the target of, I would say, 873 00:45:45,290 --> 00:45:50,920 a wonder drug-- the wonder drugs of the statins. 874 00:45:50,920 --> 00:45:53,890 So people really care about the detailed mechanism. 875 00:45:53,890 --> 00:45:56,560 We don't care about the detailed mechanism. 876 00:45:56,560 --> 00:45:59,950 We do care that hydride attacks the carbonyl, 877 00:45:59,950 --> 00:46:02,650 and it attacks the carbon and not the oxygen. 878 00:46:02,650 --> 00:46:04,810 But the details, if you're interested in that, 879 00:46:04,810 --> 00:46:07,540 you can go read about this in the reference. 880 00:46:07,540 --> 00:46:09,550 A lot of people have focused a lot of energy 881 00:46:09,550 --> 00:46:14,050 on this, trying to make better statin inhibitors. 882 00:46:14,050 --> 00:46:15,910 So what do we know about this? 883 00:46:15,910 --> 00:46:18,670 And there's a few things I want to say about this. 884 00:46:18,670 --> 00:46:20,770 And so if we look at the protein, 885 00:46:20,770 --> 00:46:26,620 we're going to come back to this in lecture 3. 886 00:46:26,620 --> 00:46:29,790 So this is important to remember. 887 00:46:29,790 --> 00:46:31,280 So this is the protein. 888 00:46:31,280 --> 00:46:33,370 I'm going to use this cartoon. 889 00:46:33,370 --> 00:46:36,090 And Liz used these cartoons as well. 890 00:46:36,090 --> 00:46:39,770 But what we're going to see is the protein 891 00:46:39,770 --> 00:46:42,120 has eight of these things-- 892 00:46:42,120 --> 00:46:43,217 eight. 893 00:46:43,217 --> 00:46:44,300 Each one of these things-- 894 00:46:48,280 --> 00:46:51,030 OK, [INAUDIBLE],, and we need three more. 895 00:46:51,030 --> 00:46:52,930 I'm not going to fit this. 896 00:46:52,930 --> 00:46:55,440 So it has a transmembrane helices. 897 00:47:00,860 --> 00:47:06,545 And the protein itself is, again, 888 amino acids. 898 00:47:09,640 --> 00:47:11,410 And what's interesting about this, 899 00:47:11,410 --> 00:47:15,070 if you have this many transmembrane helices, where's 900 00:47:15,070 --> 00:47:16,650 the protein going to be located? 901 00:47:16,650 --> 00:47:19,150 It's going to be located in a membrane. 902 00:47:19,150 --> 00:47:28,380 So these five are called the sterile sensor domain. 903 00:47:28,380 --> 00:47:33,660 HMG-CoA reductase is going to be a key player in regulation 904 00:47:33,660 --> 00:47:35,670 of cholesterol levels. 905 00:47:35,670 --> 00:47:39,780 And it exists-- it's found, this protein 906 00:47:39,780 --> 00:47:41,400 is found in the ER membrane. 907 00:47:46,260 --> 00:47:56,950 And SSD is the sterile sensor domain. 908 00:47:56,950 --> 00:47:59,070 And we're to come back to this when we start 909 00:47:59,070 --> 00:48:00,390 talking about homeostasis. 910 00:48:00,390 --> 00:48:04,050 We're going to see that there are other proteins that also 911 00:48:04,050 --> 00:48:08,130 have transmembrane helices that somehow bind and sense 912 00:48:08,130 --> 00:48:09,870 cholesterol that are going to help us 913 00:48:09,870 --> 00:48:12,360 control cholesterol levels. 914 00:48:12,360 --> 00:48:14,580 Now, what's really interesting about this-- 915 00:48:14,580 --> 00:48:16,130 so the protein is huge. 916 00:48:16,130 --> 00:48:17,730 It's stuck in membrane. 917 00:48:17,730 --> 00:48:19,740 What's really interesting is that you 918 00:48:19,740 --> 00:48:22,530 can cut the protein in half, about in half. 919 00:48:22,530 --> 00:48:24,030 That's what you're looking at there. 920 00:48:24,030 --> 00:48:25,488 You have a soluble protein, they're 921 00:48:25,488 --> 00:48:28,470 much easier to crystallize than membrane proteins. 922 00:48:28,470 --> 00:48:33,150 And it turns out, if you cut the protein in half, this guy, 923 00:48:33,150 --> 00:48:38,062 if you cut, is active. 924 00:48:38,062 --> 00:48:38,770 And it's soluble. 925 00:48:41,540 --> 00:48:44,660 And the activity is the same as the protein 926 00:48:44,660 --> 00:48:45,920 bound to the membrane. 927 00:48:45,920 --> 00:48:47,450 So it has very high activity. 928 00:48:47,450 --> 00:48:49,910 So it's like you have two separate domains. 929 00:48:49,910 --> 00:48:52,850 Furthermore, if you cut this in half, 930 00:48:52,850 --> 00:48:55,910 you can still target this to the ER membrane, 931 00:48:55,910 --> 00:48:58,470 and you can still sense cholesterol. 932 00:48:58,470 --> 00:49:00,950 So somehow these two things have come together. 933 00:49:00,950 --> 00:49:03,110 They have two really independent activities. 934 00:49:03,110 --> 00:49:05,060 But we're going to see, they work together 935 00:49:05,060 --> 00:49:08,540 to control cholesterol levels. 936 00:49:08,540 --> 00:49:09,930 So what I want to do-- 937 00:49:09,930 --> 00:49:10,770 how am I doing? 938 00:49:10,770 --> 00:49:14,000 Oh, see, time goes by too fast. 939 00:49:14,000 --> 00:49:15,890 Isn't time going by too fast for you? 940 00:49:15,890 --> 00:49:18,880 Anyhow, I want to show you-- 941 00:49:18,880 --> 00:49:20,840 and we'll come back to it next time-- 942 00:49:20,840 --> 00:49:27,080 is that the statins are the target of HMG-CoA reductase. 943 00:49:27,080 --> 00:49:29,300 I mean, this is like an amazing thing. 944 00:49:29,300 --> 00:49:34,850 Cholesterol biosynthesis was only elucidated in 1955. 945 00:49:34,850 --> 00:49:38,660 And it turns out this guy, Endo, was the first one 946 00:49:38,660 --> 00:49:42,680 to discover a natural product that somehow could 947 00:49:42,680 --> 00:49:45,770 lower cholesterol in 1976. 948 00:49:45,770 --> 00:49:49,040 And actually, when I was a young person, 949 00:49:49,040 --> 00:49:50,930 Al Alberts used to work at Merck. 950 00:49:50,930 --> 00:49:53,060 I used to consult for Merck back in those days. 951 00:49:53,060 --> 00:49:55,850 It was incredibly exciting times, 952 00:49:55,850 --> 00:49:58,880 because he discovered really sort 953 00:49:58,880 --> 00:50:02,490 of the first real statin that worked, that wasn't toxic-- 954 00:50:02,490 --> 00:50:04,250 lovastatin. 955 00:50:04,250 --> 00:50:07,070 And really, within a period of only seven years, 956 00:50:07,070 --> 00:50:08,870 this was approved by the FDA. 957 00:50:08,870 --> 00:50:10,580 So that's an amazing observation. 958 00:50:10,580 --> 00:50:14,090 People are still gobbling down statins everywhere. 959 00:50:14,090 --> 00:50:18,596 There are issues with them, but it makes $30 billion 960 00:50:18,596 --> 00:50:21,440 for the companies that own this. 961 00:50:21,440 --> 00:50:26,110 So you now might have heard of Lipitor or Crestor-- 962 00:50:26,110 --> 00:50:29,010 anyhow, they are there. 963 00:50:29,010 --> 00:50:31,340 And it really is a wonder drug. 964 00:50:31,340 --> 00:50:34,760 And it works, we're going to see next time. 965 00:50:34,760 --> 00:50:37,400 Because it looks like the substrate 966 00:50:37,400 --> 00:50:41,150 hydroxymethylglutaryl-CoA-- 967 00:50:41,150 --> 00:50:44,180 So that it acts as a competitive inhibitor 968 00:50:44,180 --> 00:50:48,620 for binding to the active site of HMG-CoA reductase, 969 00:50:48,620 --> 00:50:51,200 and prevents the reduction process. 970 00:50:51,200 --> 00:50:53,330 And we'll come back next time and look 971 00:50:53,330 --> 00:50:55,157 at a little bit at the details. 972 00:50:55,157 --> 00:50:57,740 We're not going to spend a lot of time looking at the details, 973 00:50:57,740 --> 00:51:02,300 but then finish on to get to IPP and dimethyl APP, 974 00:51:02,300 --> 00:51:06,550 the building blocks we're after to make all terpenes. 975 00:51:06,550 --> 00:51:08,100 OK.