1 00:00:00,500 --> 00:00:02,820 The following content is provided under a Creative 2 00:00:02,820 --> 00:00:04,360 Commons license. 3 00:00:04,360 --> 00:00:06,660 Your support will help MIT OpenCourseWare 4 00:00:06,660 --> 00:00:11,020 continue to offer high quality educational resources for free. 5 00:00:11,020 --> 00:00:13,650 To make a donation or view additional materials 6 00:00:13,650 --> 00:00:17,600 from hundreds of MIT courses, visit MIT OpenCourseWare 7 00:00:17,600 --> 00:00:18,540 at ocw.mit.edu. 8 00:00:25,270 --> 00:00:26,770 JOANNE STUBBE: So we've been talking 9 00:00:26,770 --> 00:00:32,570 about iron metabolism in general in the first lecture. 10 00:00:32,570 --> 00:00:34,480 And in the second lecture we started 11 00:00:34,480 --> 00:00:37,090 to focus on iron metabolism in humans, 12 00:00:37,090 --> 00:00:39,190 and the third set of lectures is going 13 00:00:39,190 --> 00:00:45,020 to be iron metabolism and bacteria with a focus on hemes. 14 00:00:45,020 --> 00:00:49,000 And the two things you want to talk about in the lecture today 15 00:00:49,000 --> 00:00:53,900 are, how does iron get taken up into cells in humans, 16 00:00:53,900 --> 00:00:57,670 with a focus on receptor mediated endocytosis, 17 00:00:57,670 --> 00:01:00,280 and then we're going to start talking about hopefully iron 18 00:01:00,280 --> 00:01:02,020 regulation-- 19 00:01:02,020 --> 00:01:05,500 how you sense iron, ion regulation 20 00:01:05,500 --> 00:01:09,130 at the translational level. 21 00:01:09,130 --> 00:01:13,540 By sort of a unique mechanism, at least 22 00:01:13,540 --> 00:01:15,940 at the time of its discovery. 23 00:01:15,940 --> 00:01:19,960 So in the last lecture, we introduced you 24 00:01:19,960 --> 00:01:22,690 to some key features about iron chemistry 25 00:01:22,690 --> 00:01:25,780 in general that we're going to use throughout this lecture 26 00:01:25,780 --> 00:01:26,700 and next lecture. 27 00:01:26,700 --> 00:01:28,615 So you need to go back and review your notes 28 00:01:28,615 --> 00:01:29,740 if you don't remember that. 29 00:01:29,740 --> 00:01:32,200 Or hopefully you've had it somewhere before, 30 00:01:32,200 --> 00:01:35,170 and it's a review from you from freshman chemistry 31 00:01:35,170 --> 00:01:37,870 or inorganic chemistry. 32 00:01:37,870 --> 00:01:40,745 And so iron metabolism-- what do we know? 33 00:01:40,745 --> 00:01:45,807 We know the average human being has 3 to 4 grams of iron. 34 00:01:45,807 --> 00:01:47,890 We talked about this at the end of the last class, 35 00:01:47,890 --> 00:01:50,110 of how is the iron distributed. 36 00:01:50,110 --> 00:01:52,600 We all went through that most of our iron 37 00:01:52,600 --> 00:01:57,410 is in our red blood cells in the form of hemoglobin. 38 00:01:57,410 --> 00:02:01,180 But it's also-- so in the form of hemoglobin, 39 00:02:01,180 --> 00:02:06,280 it also can be stored in proteins called ferritins, 40 00:02:06,280 --> 00:02:08,199 which we're not going to spend much time on, 41 00:02:08,199 --> 00:02:11,440 but I will introduce you to today. 42 00:02:11,440 --> 00:02:14,740 And then many of you may know that red blood 43 00:02:14,740 --> 00:02:18,190 cells die every 120 days. 44 00:02:18,190 --> 00:02:22,960 And we'll see that the iron is really continually recycled, 45 00:02:22,960 --> 00:02:25,660 and we'll talk a little bit about the mechanism 46 00:02:25,660 --> 00:02:27,100 of how that's regulated. 47 00:02:27,100 --> 00:02:30,880 So instead of excreting it, what happens is you recycle. 48 00:02:30,880 --> 00:02:34,990 The iron unit's recycled by macrophages in the spleen. 49 00:02:34,990 --> 00:02:38,650 And so the other place you see a fair amount of iron 50 00:02:38,650 --> 00:02:40,280 is in the macrophages. 51 00:02:40,280 --> 00:02:42,760 And the third place you see a fair amount of iron 52 00:02:42,760 --> 00:02:45,700 is in the tissues, because myoglobin, again, 53 00:02:45,700 --> 00:02:49,750 has to deliver oxygen to the respiratory chain. 54 00:02:49,750 --> 00:02:52,480 So what I want to do now, and I'm 55 00:02:52,480 --> 00:02:57,670 going to go back and forth between the PowerPoint 56 00:02:57,670 --> 00:02:59,230 and notes. 57 00:02:59,230 --> 00:03:01,810 And so some things I'm going to write down some things not. 58 00:03:01,810 --> 00:03:04,313 Hopefully you have these cartoons in front of you 59 00:03:04,313 --> 00:03:05,980 so you can write down some of the things 60 00:03:05,980 --> 00:03:10,120 that I will say here, and say it again and say it again. 61 00:03:10,120 --> 00:03:13,540 So this is sort of the big picture 62 00:03:13,540 --> 00:03:14,860 that I took from some review. 63 00:03:14,860 --> 00:03:17,832 And most of these big pictures have some issues with them. 64 00:03:17,832 --> 00:03:19,790 But I think it still gives you the big picture. 65 00:03:19,790 --> 00:03:25,600 So here's a duodenum, where we can take up iron from the diet. 66 00:03:25,600 --> 00:03:27,760 And we'll talk about this in more detail, 67 00:03:27,760 --> 00:03:33,130 but a key player in allowing the iron from the diet 68 00:03:33,130 --> 00:03:36,490 to go into our system is going to be FPN-- 69 00:03:36,490 --> 00:03:38,110 that's going to be ferroportin, I'm 70 00:03:38,110 --> 00:03:39,760 going to describe this again. 71 00:03:39,760 --> 00:03:42,680 But you're going to see FPN over and over again. 72 00:03:42,680 --> 00:03:47,830 It allows iron to be transferred in the plus 2 state, 73 00:03:47,830 --> 00:03:50,390 and that's going to be important. 74 00:03:50,390 --> 00:03:53,490 And so what we see, that if you look at iron from the diet, 75 00:03:53,490 --> 00:03:55,300 there's not that much. 76 00:03:55,300 --> 00:03:58,780 [AUDIO OUT] somebody's guess as to how much there is. 77 00:03:58,780 --> 00:03:59,860 A few milligrams. 78 00:03:59,860 --> 00:04:03,250 And the question is, where does it go in the bloodstream? 79 00:04:03,250 --> 00:04:05,620 And it goes to a protein that we're 80 00:04:05,620 --> 00:04:11,050 going to talk about that's a carrier for iron in the plus 3 81 00:04:11,050 --> 00:04:11,560 state. 82 00:04:11,560 --> 00:04:15,580 So we're going to see plus 2, plus 3 into conversions 83 00:04:15,580 --> 00:04:16,519 over and over again. 84 00:04:16,519 --> 00:04:19,000 And sort of what the strategy that 85 00:04:19,000 --> 00:04:23,800 has evolved to be able to deal with these different oxidation 86 00:04:23,800 --> 00:04:24,640 states is. 87 00:04:24,640 --> 00:04:28,787 We'll see that this little protein, TF, is transferrin, 88 00:04:28,787 --> 00:04:30,370 and we're going to look at transferrin 89 00:04:30,370 --> 00:04:36,100 for a-- very briefly, but it binds iron 3 and bicarbonate, 90 00:04:36,100 --> 00:04:41,230 and then delivers this to tissues, 91 00:04:41,230 --> 00:04:44,770 and also delivers it to marrow. 92 00:04:44,770 --> 00:04:48,310 And marrow, which is-- accounts for approximately, 93 00:04:48,310 --> 00:04:52,450 by mass, 4% of the body weight, makes all of our red and white 94 00:04:52,450 --> 00:04:53,900 blood cells. 95 00:04:53,900 --> 00:04:55,880 So that's going to be important. 96 00:04:55,880 --> 00:05:01,420 And so the marrow makes the erythrocyte, 97 00:05:01,420 --> 00:05:05,020 the heme for the erythrocytes makes the erythrocytes, 98 00:05:05,020 --> 00:05:07,240 and the erythrocytes are the red blood cells 99 00:05:07,240 --> 00:05:09,590 that have all the hemoglobin. 100 00:05:09,590 --> 00:05:14,890 So out of the 4 grams, you have 2 and 1/2 grams of hemoglobin. 101 00:05:14,890 --> 00:05:19,960 And then these red blood cells die every 120 days, 102 00:05:19,960 --> 00:05:23,410 and instead of just discarding everything, they're recycled. 103 00:05:23,410 --> 00:05:27,820 And they're recycled by the macrophages in the spleen. 104 00:05:27,820 --> 00:05:32,800 And somehow you want to take the iron from these red blood cells 105 00:05:32,800 --> 00:05:34,510 and reuse it. 106 00:05:34,510 --> 00:05:38,290 And so there's a series of reactions that happen. 107 00:05:38,290 --> 00:05:42,280 Ultimately you get iron 2, and the iron 2-- here's 108 00:05:42,280 --> 00:05:47,020 again our iron 2 transporter, ferroportin, is 109 00:05:47,020 --> 00:05:51,250 going to take the iron that's recovered and put it 110 00:05:51,250 --> 00:05:54,760 back into transferrin, where, again, it can be distributed, 111 00:05:54,760 --> 00:05:57,790 depending on the sensing of iron. 112 00:05:57,790 --> 00:06:02,890 Now, the major player in the sensing and storage of iron 113 00:06:02,890 --> 00:06:04,263 is the liver. 114 00:06:04,263 --> 00:06:05,930 So the liver, we're going to see there's 115 00:06:05,930 --> 00:06:09,190 a protein there not indicated on the slide called ferritin, 116 00:06:09,190 --> 00:06:16,060 and ferritin binds 4,500 molecules of iron. 117 00:06:16,060 --> 00:06:23,440 And this is also-- the liver is the organ that generates, 118 00:06:23,440 --> 00:06:27,950 biosynthesizes the key regulator of iron homeostasis, 119 00:06:27,950 --> 00:06:29,933 which is a peptide hormone that we're not 120 00:06:29,933 --> 00:06:31,600 going to spend a lot of time on, but I'm 121 00:06:31,600 --> 00:06:33,640 going to show you what it does. 122 00:06:33,640 --> 00:06:35,800 So that's called hepcidin. 123 00:06:35,800 --> 00:06:41,020 And what we'll see is hepcidin in some way controls 124 00:06:41,020 --> 00:06:44,140 the levels of ferroportin. 125 00:06:44,140 --> 00:06:48,400 So we also see that we lose some iron daily, 126 00:06:48,400 --> 00:06:51,130 but the iron losses are small. 127 00:06:51,130 --> 00:06:53,440 So we have a lot of iron units, but the iron 128 00:06:53,440 --> 00:06:55,630 is continually recycled, and the question 129 00:06:55,630 --> 00:06:58,960 is, how does that happen? 130 00:06:58,960 --> 00:07:04,860 So I just want to look at one place where, 131 00:07:04,860 --> 00:07:08,200 in the duodenum, where we're going to take up iron. 132 00:07:08,200 --> 00:07:10,690 So what I'm going to do is-- 133 00:07:10,690 --> 00:07:13,610 this is a cartoon of what I just showed you in more detail. 134 00:07:13,610 --> 00:07:19,170 But I'm going to focus on iron absorption from the diet. 135 00:07:19,170 --> 00:07:20,710 And I want to make a couple points 136 00:07:20,710 --> 00:07:23,290 about this, which are general. 137 00:07:23,290 --> 00:07:28,000 And so what we'll see is we have enterocytes, 138 00:07:28,000 --> 00:07:29,560 so this is an enterocyte. 139 00:07:34,650 --> 00:07:37,500 And you have an apical brush border membrane. 140 00:07:48,240 --> 00:07:50,580 And then you have a second membrane 141 00:07:50,580 --> 00:07:52,720 which is going to get us into the bloodstream. 142 00:07:52,720 --> 00:07:54,870 So this is called a basolateral membrane. 143 00:08:02,070 --> 00:08:07,200 So we get iron from our diets mostly in the plus 3 state. 144 00:08:07,200 --> 00:08:09,780 But to do anything with iron, probably 145 00:08:09,780 --> 00:08:11,820 because of the ligand exchange issues 146 00:08:11,820 --> 00:08:15,650 we talked about last time, the rate constants for exchange 147 00:08:15,650 --> 00:08:18,450 are much slower with iron 3 than iron 2. 148 00:08:18,450 --> 00:08:21,915 So from the diet, we have iron 3. 149 00:08:24,610 --> 00:08:31,833 And iron 3 needs to be reduced to iron 2. 150 00:08:31,833 --> 00:08:33,750 And that can be done-- we'll see this is going 151 00:08:33,750 --> 00:08:36,559 to happen over and over again. 152 00:08:36,559 --> 00:08:41,230 And this can be done by a ferric reductase. 153 00:08:46,370 --> 00:08:52,570 And what we will see is in this membrane, 154 00:08:52,570 --> 00:08:55,250 we're going to have an iron 2 transporter. 155 00:08:55,250 --> 00:08:58,960 So in addition to the ferroportin 156 00:08:58,960 --> 00:09:00,950 I just briefly introduced you to, 157 00:09:00,950 --> 00:09:04,160 and will introduce you to again, we 158 00:09:04,160 --> 00:09:08,570 have an iron 2 transporter, that's called DMT 1. 159 00:09:08,570 --> 00:09:11,630 Again, the acronyms are horrible. 160 00:09:11,630 --> 00:09:15,560 But it's a divalent predominantly iron 161 00:09:15,560 --> 00:09:17,090 2 metal transporter. 162 00:09:24,048 --> 00:09:25,590 And we're going to see, when we think 163 00:09:25,590 --> 00:09:29,640 about regulation of iron homeostasis, 164 00:09:29,640 --> 00:09:31,320 this is going to be a key player. 165 00:09:31,320 --> 00:09:35,730 Because it takes iron from the diet into our cells. 166 00:09:35,730 --> 00:09:39,930 And in this membrane of the enterocyte, what 167 00:09:39,930 --> 00:09:43,730 we will see is that we have-- 168 00:09:43,730 --> 00:09:47,460 and this is what you saw in the previous slide-- 169 00:09:47,460 --> 00:09:52,740 you have ferroportin-- so I'm only 170 00:09:52,740 --> 00:09:54,660 going to write this down once. 171 00:09:57,510 --> 00:10:02,790 But this is going to take the iron 2 172 00:10:02,790 --> 00:10:07,110 and then transfer it into, ultimately, 173 00:10:07,110 --> 00:10:08,970 the carrier in the bloodstream, which 174 00:10:08,970 --> 00:10:12,920 is going to be transferrin. 175 00:10:12,920 --> 00:10:16,650 So here we have iron 2, but for it 176 00:10:16,650 --> 00:10:21,382 to get picked up by transferrin, it gets oxidized to iron 3. 177 00:10:21,382 --> 00:10:23,340 So what you're going to see over and over again 178 00:10:23,340 --> 00:10:28,290 is going back and forth between iron 2 and iron 3. 179 00:10:28,290 --> 00:10:33,270 And so this gets oxidized to iron 3. 180 00:10:33,270 --> 00:10:40,085 And these proteins-- there's a copper iron oxidase. 181 00:10:42,695 --> 00:10:44,320 And if you look at the handouts, you'll 182 00:10:44,320 --> 00:10:46,700 see that this is also called-- 183 00:10:52,170 --> 00:10:54,320 again, I don't expect you remember the names. 184 00:10:54,320 --> 00:11:00,560 What I think is key here is that you need to transfer this 185 00:11:00,560 --> 00:11:04,340 to the plus 3 oxidation state. 186 00:11:04,340 --> 00:11:08,440 So now what happens in the plus 3 oxidation state-- 187 00:11:08,440 --> 00:11:12,260 so let's go over to the next board here-- 188 00:11:12,260 --> 00:11:16,920 we have a protein called transferrin, 189 00:11:16,920 --> 00:11:20,920 and we'll look at this a little bit. 190 00:11:20,920 --> 00:11:25,480 And transferrin is going to bind iron in the plus 3 state, 191 00:11:25,480 --> 00:11:28,750 but it also requires bicarbonate. 192 00:11:28,750 --> 00:11:31,630 So in the blood, is that unusual that you 193 00:11:31,630 --> 00:11:33,910 would require bicarbonate? 194 00:11:33,910 --> 00:11:36,520 Or why might you require bicarbonate? 195 00:11:36,520 --> 00:11:40,180 What do you know about blood cells and hemoglobin? 196 00:11:44,380 --> 00:11:49,330 So we have iron 3 that's regenerated enzymatically, 197 00:11:49,330 --> 00:11:51,477 through some kind of oxidation reduction equipment. 198 00:11:51,477 --> 00:11:53,810 And we're going to see this, again, over and over again. 199 00:11:53,810 --> 00:11:55,268 And they each have different names, 200 00:11:55,268 --> 00:11:56,890 so that's confusing as well. 201 00:11:56,890 --> 00:11:59,980 But you're cycling between 2 and 3. 202 00:11:59,980 --> 00:12:01,750 And then transferrin, we have a structure 203 00:12:01,750 --> 00:12:05,500 of this picks up the iron in the plus 3 state, 204 00:12:05,500 --> 00:12:07,457 and also picks up bicarbonate. 205 00:12:07,457 --> 00:12:09,040 So where do you think that bicarbonate 206 00:12:09,040 --> 00:12:10,970 comes from in blood cells? 207 00:12:10,970 --> 00:12:11,850 AUDIENCE: CO2. 208 00:12:11,850 --> 00:12:13,270 JOANNE STUBBE: Yeah, so it comes from CO2. 209 00:12:13,270 --> 00:12:13,780 Why? 210 00:12:13,780 --> 00:12:15,340 Because a major function of red blood 211 00:12:15,340 --> 00:12:19,870 cells is to transfer CO2 from the tissues back to the lungs. 212 00:12:19,870 --> 00:12:27,280 So CO2 is not there, at pH 7, it gets rapidly hydrated to form 213 00:12:27,280 --> 00:12:30,070 bicarbonate and protons. 214 00:12:30,070 --> 00:12:32,658 And so this is unusual. 215 00:12:32,658 --> 00:12:35,200 I think this is one of the few systems where you have-- we'll 216 00:12:35,200 --> 00:12:38,230 see bicarbonate as a ligand. 217 00:12:38,230 --> 00:12:43,240 So in addition to these enterocytes, which again 218 00:12:43,240 --> 00:12:47,890 are involved in iron uptake, we also 219 00:12:47,890 --> 00:12:52,385 have macrophages in the spleen. 220 00:12:58,750 --> 00:13:01,165 And so this, again, is due to the diet. 221 00:13:06,580 --> 00:13:12,160 And this is due to basically recycling-- 222 00:13:12,160 --> 00:13:14,470 iron recycling. 223 00:13:14,470 --> 00:13:17,920 And so what you have is macrophages in the spleen, 224 00:13:17,920 --> 00:13:25,240 and you have in the macrophages dead red blood cells, 225 00:13:25,240 --> 00:13:28,210 which I'll abbreviate RBC. 226 00:13:28,210 --> 00:13:30,790 And so the idea is we want to get the iron out 227 00:13:30,790 --> 00:13:34,960 of the red blood cells somehow to reuse it. 228 00:13:34,960 --> 00:13:37,180 So that's the goal. 229 00:13:37,180 --> 00:13:45,520 And so somehow in a complicated process, we get iron 2 out. 230 00:13:45,520 --> 00:13:46,705 And then iron 2-- 231 00:13:50,200 --> 00:13:52,930 here we have our friend ferroportin, 232 00:13:52,930 --> 00:13:56,770 that I just showed you in the previous slide, 233 00:13:56,770 --> 00:14:02,430 is going to take and put into the extracellular mirror 234 00:14:02,430 --> 00:14:05,670 in the plasma the iron 2. 235 00:14:05,670 --> 00:14:07,670 So what happens to the iron 2? 236 00:14:07,670 --> 00:14:12,870 We just saw over here, the iron 2 gets oxidized to iron 3. 237 00:14:12,870 --> 00:14:15,400 The same thing is going to happen over here. 238 00:14:15,400 --> 00:14:23,730 So we have iron 2 that needs to get oxidized to iron 3. 239 00:14:23,730 --> 00:14:28,980 And again, let's just call it a copper iron oxidase. 240 00:14:28,980 --> 00:14:30,990 I'm not going to go through the details. 241 00:14:30,990 --> 00:14:33,850 And then what happens to the iron 3? 242 00:14:33,850 --> 00:14:37,400 So the iron 3 then gets picked up by the transferrin. 243 00:14:37,400 --> 00:14:40,920 And then depending on what the needs are the cell, 244 00:14:40,920 --> 00:14:44,220 the transferrin can deliver. 245 00:14:44,220 --> 00:14:46,590 If you have a lot of iron, it could deliver it back 246 00:14:46,590 --> 00:14:47,280 to the liver. 247 00:14:47,280 --> 00:14:50,040 We'll see that's the storage place for the iron. 248 00:14:50,040 --> 00:14:52,740 So the iron 3 transferrin needs to get taken up, 249 00:14:52,740 --> 00:14:55,920 just like we saw with cholesterol. 250 00:14:55,920 --> 00:15:00,210 Or if we need iron in some other tissues, 251 00:15:00,210 --> 00:15:03,270 we'll see that there are receptors for iron 3 252 00:15:03,270 --> 00:15:06,150 transferrin that can, again, take iron into the cells 253 00:15:06,150 --> 00:15:10,440 to meet the needs of the cell for iron requirement. 254 00:15:10,440 --> 00:15:13,200 Now, the one thing I wanted to tell you in the first slide, 255 00:15:13,200 --> 00:15:17,220 which I had forgot, was that in addition 256 00:15:17,220 --> 00:15:20,130 to all of these requirements for iron, 257 00:15:20,130 --> 00:15:25,470 and the predominant form being hemoglobin and myoglobin, what 258 00:15:25,470 --> 00:15:28,470 we see is that iron is found in only 4% 259 00:15:28,470 --> 00:15:30,780 of the metabolic enzymes. 260 00:15:30,780 --> 00:15:33,000 So iron is found in many proteins that 261 00:15:33,000 --> 00:15:35,730 catalyze all kinds of reactions, like we talked about last time. 262 00:15:35,730 --> 00:15:39,660 But that's a small percentage of the total amount of iron. 263 00:15:43,710 --> 00:15:47,310 So this sort of diagram is pointing out 264 00:15:47,310 --> 00:15:51,270 a few things that sort of is indicative of iron 265 00:15:51,270 --> 00:15:54,840 mediated metabolism in many cases. 266 00:15:54,840 --> 00:15:57,360 And so what I briefly want to do is 267 00:15:57,360 --> 00:16:02,800 sort of summarize the functions of these different proteins. 268 00:16:02,800 --> 00:16:04,260 So this is phenomenological. 269 00:16:04,260 --> 00:16:06,360 And if you're going to have-- 270 00:16:06,360 --> 00:16:07,920 if you were given an exam on this, 271 00:16:07,920 --> 00:16:11,130 I'll give you the names of all of these things. 272 00:16:11,130 --> 00:16:13,680 Because I think the names are actually confusing. 273 00:16:13,680 --> 00:16:19,620 So number one, we have DMT1. 274 00:16:19,620 --> 00:16:22,690 And again, when ion is trans-- 275 00:16:22,690 --> 00:16:25,560 it's a transporter of iron 2. 276 00:16:31,550 --> 00:16:34,500 And so that's an important thing to remember. 277 00:16:34,500 --> 00:16:37,430 But even though it's transferred into the cell 278 00:16:37,430 --> 00:16:42,290 and it moves around inside the cell as iron 2, likely because, 279 00:16:42,290 --> 00:16:45,330 again, the ligand exchange, though iron starts here 280 00:16:45,330 --> 00:16:48,360 and it needs to move here, and it needs to move here, 281 00:16:48,360 --> 00:16:50,760 and this is the way nature-- 282 00:16:50,760 --> 00:16:53,520 because of the exchangeability of the ligands-- 283 00:16:53,520 --> 00:16:56,130 has decided to move iron, and also 284 00:16:56,130 --> 00:16:58,230 oftentimes copper 1 around, instead 285 00:16:58,230 --> 00:17:04,829 of in the oxidized state that this transporter deals 286 00:17:04,829 --> 00:17:06,810 with iron 2. 287 00:17:06,810 --> 00:17:12,060 The second key thing is ferroportin, and this goes-- 288 00:17:12,060 --> 00:17:22,609 brings, again, iron 2 to the extracellular milieu. 289 00:17:25,130 --> 00:17:28,084 And so this is bringing it inside the cell. 290 00:17:33,670 --> 00:17:35,790 This is bringing it extracellularly 291 00:17:35,790 --> 00:17:37,760 the outside the cell. 292 00:17:37,760 --> 00:17:39,940 And this leads to the next thing that we 293 00:17:39,940 --> 00:17:41,590 see over and over again-- 294 00:17:41,590 --> 00:17:44,430 while iron 2 is brought outside the cell, 295 00:17:44,430 --> 00:17:47,440 it then gets oxidized to do anything with it. 296 00:17:47,440 --> 00:17:53,950 So then we have general ways of iron 2 297 00:17:53,950 --> 00:17:57,620 being oxidized to iron 3. 298 00:17:57,620 --> 00:18:03,580 And this could be a copper iron oxidase. 299 00:18:03,580 --> 00:18:05,970 But again, there are multiple-- 300 00:18:05,970 --> 00:18:08,460 there are multiple names for these [INAUDIBLE]---- 301 00:18:08,460 --> 00:18:10,210 we'll see in a few minutes, steep is one. 302 00:18:10,210 --> 00:18:15,280 I mean, they have five different iron oxidases. 303 00:18:15,280 --> 00:18:21,130 And iron 3 is going to be the key for transferring this 304 00:18:21,130 --> 00:18:25,450 to ferritin, which is the way that iron is transferred, 305 00:18:25,450 --> 00:18:29,170 just like the LDL particles are the way cholesterol is 306 00:18:29,170 --> 00:18:31,520 transferred around the cell. 307 00:18:31,520 --> 00:18:34,870 Ferritin-- transferrin is the way 308 00:18:34,870 --> 00:18:38,000 the iron is transferred around the cell. 309 00:18:38,000 --> 00:18:43,690 So so this iron 3-- 310 00:18:43,690 --> 00:18:48,820 so iron 3 is picked up by transferrin. 311 00:18:55,680 --> 00:19:02,160 And again, this is iron 3. 312 00:19:02,160 --> 00:19:03,630 I'll show you-- we have structures 313 00:19:03,630 --> 00:19:05,370 of all these proteins. 314 00:19:05,370 --> 00:19:09,450 This is, again, iron 3 bicarbonate. 315 00:19:09,450 --> 00:19:13,200 And then the question is, how does this transferrin 316 00:19:13,200 --> 00:19:14,760 get into cells? 317 00:19:14,760 --> 00:19:16,150 So this is the major carrier. 318 00:19:21,350 --> 00:19:21,850 Iron. 319 00:19:25,780 --> 00:19:29,370 And it's carried in the plus 3 oxidation state. 320 00:19:29,370 --> 00:19:32,050 Maybe, and we'll see that the KD for binding-- 321 00:19:32,050 --> 00:19:34,660 what do you think the KD for binding to a transferrin 322 00:19:34,660 --> 00:19:35,160 might be? 323 00:19:35,160 --> 00:19:36,118 Do you think it's weak? 324 00:19:36,118 --> 00:19:37,740 Do you think it's strong? 325 00:19:37,740 --> 00:19:41,190 And what would you, if you were designing something 326 00:19:41,190 --> 00:19:43,263 that was carrying around iron to all the tissues, 327 00:19:43,263 --> 00:19:44,180 what would you design? 328 00:19:46,690 --> 00:19:47,830 Something weak or strong? 329 00:19:54,940 --> 00:19:56,530 Say it was weak, what would happen? 330 00:20:02,695 --> 00:20:04,680 Yeah, it comes unbound. 331 00:20:04,680 --> 00:20:07,245 And then if it gets reduced, into the realm where 332 00:20:07,245 --> 00:20:11,160 you have iron 2 and then you have reactive oxygen species. 333 00:20:11,160 --> 00:20:14,550 And so nature has developed, I would say, 334 00:20:14,550 --> 00:20:16,110 you've seen with siderophores you 335 00:20:16,110 --> 00:20:19,710 can get things 10 to the minus 35 for dissociation constants 336 00:20:19,710 --> 00:20:21,400 to 10 to the minus 50. 337 00:20:21,400 --> 00:20:27,210 The KD for this is 10 to the minus 23 molar 338 00:20:27,210 --> 00:20:30,210 for iron binding to transferrin. 339 00:20:30,210 --> 00:20:33,840 And so the next thing that happens 340 00:20:33,840 --> 00:20:39,300 is that the iron binding to transference 341 00:20:39,300 --> 00:20:42,910 goes to the transferrin receptor. 342 00:20:42,910 --> 00:20:53,760 And so transferrin then binds to the transferrin receptor, 343 00:20:53,760 --> 00:20:58,770 just like the LDL particle binds to the LDL receptor. 344 00:20:58,770 --> 00:21:06,575 So this is the transferrin receptor. 345 00:21:09,330 --> 00:21:12,110 And so what you're going to see is that, in contrast 346 00:21:12,110 --> 00:21:14,720 with iron transported across-- 347 00:21:14,720 --> 00:21:18,410 in the case of the enterocyte, or in the case of ferroportin, 348 00:21:18,410 --> 00:21:22,520 where it's iron 2, this is all transferred in the iron 3 349 00:21:22,520 --> 00:21:23,720 state. 350 00:21:23,720 --> 00:21:28,070 So this-- again, this is important to see 351 00:21:28,070 --> 00:21:31,160 the differences in the oxidation states that 352 00:21:31,160 --> 00:21:35,580 are used to control uptake into the cell. 353 00:21:35,580 --> 00:21:38,090 And this occurs by-- 354 00:21:38,090 --> 00:21:39,800 we'll briefly look at this, but it's 355 00:21:39,800 --> 00:21:44,920 very similar to what you saw with the LDL receptor. 356 00:21:44,920 --> 00:21:47,677 The receptor mediated endocytosis. 357 00:21:47,677 --> 00:21:49,510 So we're going to look at a cartoon of this. 358 00:21:57,280 --> 00:22:01,750 So there's one other player that I want to introduce you to. 359 00:22:01,750 --> 00:22:04,270 And this player becomes really critical 360 00:22:04,270 --> 00:22:05,830 because we don't have ways-- we don't 361 00:22:05,830 --> 00:22:09,550 produce a lot of excess iron and then export it. 362 00:22:09,550 --> 00:22:11,890 All the iron is recycled. 363 00:22:11,890 --> 00:22:14,530 So what controls that iron recycling? 364 00:22:14,530 --> 00:22:28,320 So the key regulator is a peptide hormone 365 00:22:28,320 --> 00:22:32,110 which I introduced you to in the previous slide, called 366 00:22:32,110 --> 00:22:32,610 hepcidin. 367 00:22:37,900 --> 00:22:42,100 And we know quite a bit, actually, 368 00:22:42,100 --> 00:22:44,650 about the structure of this peptide hormone. 369 00:22:44,650 --> 00:22:47,110 And I'll tell you what its proposed function is. 370 00:22:47,110 --> 00:22:51,190 We're not going to spend a lot of time discussing this. 371 00:22:51,190 --> 00:22:55,240 But it is made in the liver. 372 00:22:55,240 --> 00:22:59,980 So it's bio synthesized in the liver. 373 00:23:04,330 --> 00:23:07,240 And it's basically-- its function 374 00:23:07,240 --> 00:23:09,460 is, it's a major site of regulation, 375 00:23:09,460 --> 00:23:34,630 and it controls iron from the diet, and iron cycling 376 00:23:34,630 --> 00:23:45,700 through extracellular factors, like the transferrin-- 377 00:23:45,700 --> 00:23:47,930 like transferrin. 378 00:23:47,930 --> 00:23:49,040 So how does it do this? 379 00:23:49,040 --> 00:23:51,380 So here we have a little peptide hormone. 380 00:23:51,380 --> 00:23:53,570 It's made in the liver. 381 00:23:53,570 --> 00:23:56,510 And how can a control iron recycling? 382 00:23:56,510 --> 00:24:00,200 And so the one guy that we see now 383 00:24:00,200 --> 00:24:03,090 is ferroportin, ferroportin. 384 00:24:03,090 --> 00:24:05,910 And so its major function-- it has a lot of functions, 385 00:24:05,910 --> 00:24:08,480 and it's complicated, and people are still studying this. 386 00:24:08,480 --> 00:24:10,670 But one of the major functions is 387 00:24:10,670 --> 00:24:14,720 to control the amount of ferroportin. 388 00:24:14,720 --> 00:24:18,220 So if you look at the way it's described, 389 00:24:18,220 --> 00:24:21,880 the hepcidin combined extracellularly 390 00:24:21,880 --> 00:24:23,687 to the ferroportin. 391 00:24:23,687 --> 00:24:25,270 So I'll draw a little cartoon of that. 392 00:24:31,920 --> 00:24:33,650 And then targets it for degradation 393 00:24:33,650 --> 00:24:36,890 by the proteosome inside the cell. 394 00:24:36,890 --> 00:24:39,857 So that's the key feature of hepcidin 395 00:24:39,857 --> 00:24:40,940 that you need to remember. 396 00:24:43,540 --> 00:24:45,510 So we're going to see, if you look 397 00:24:45,510 --> 00:24:49,800 at-- if you look at a lot of the cartoons I've given you, 398 00:24:49,800 --> 00:25:01,270 you have your ferroportin, watch transfers 399 00:25:01,270 --> 00:25:05,680 iron from the inside extracellularly. 400 00:25:10,480 --> 00:25:12,120 I forgot my colored chalk today. 401 00:25:12,120 --> 00:25:13,800 I was on drugs or something. 402 00:25:13,800 --> 00:25:18,780 But people were bothering me up until five minutes. 403 00:25:18,780 --> 00:25:20,970 I didn't have time to think before this lecture. 404 00:25:20,970 --> 00:25:23,640 So I'm sorry I'm a little discombobbled here. 405 00:25:23,640 --> 00:25:25,620 But this is hepcidin-- 406 00:25:28,770 --> 00:25:29,580 hep-cid-in. 407 00:25:32,290 --> 00:25:35,737 And so it binds to the extracellular side. 408 00:25:35,737 --> 00:25:37,320 And what does that does when it binds? 409 00:25:37,320 --> 00:25:39,690 It causes-- somehow things change, 410 00:25:39,690 --> 00:25:42,510 and it causes it to be degraded inside the cell 411 00:25:42,510 --> 00:25:43,570 by the proteosome. 412 00:25:43,570 --> 00:25:44,070 So. 413 00:25:44,070 --> 00:25:53,050 This interaction, extracellular, causes 414 00:25:53,050 --> 00:26:05,490 ferroportin to be degraded inside the cell 415 00:26:05,490 --> 00:26:09,130 by our friend the proteosome. 416 00:26:12,620 --> 00:26:15,500 So does everybody sort of understand what the model is? 417 00:26:15,500 --> 00:26:18,890 So this is the key regulator. 418 00:26:18,890 --> 00:26:20,780 And you've seen ferroportin-- 419 00:26:20,780 --> 00:26:22,310 we only looked at two cell types. 420 00:26:22,310 --> 00:26:30,380 We looked at the enterocyte, and we looked at the macrophages 421 00:26:30,380 --> 00:26:32,840 in the spleen, both of which have ferroportins, 422 00:26:32,840 --> 00:26:36,830 but ferroportins that are in a number of additional cell 423 00:26:36,830 --> 00:26:37,470 types. 424 00:26:37,470 --> 00:26:41,120 And when we look at regulation, one of the key regulators 425 00:26:41,120 --> 00:26:44,430 of everything is going to be that we need to control 426 00:26:44,430 --> 00:26:46,730 are the levels of ferroportin. 427 00:26:46,730 --> 00:26:50,300 Because that allows all the iron to somehow be recycled. 428 00:26:50,300 --> 00:26:52,760 It's a key player controlled by hepcidin 429 00:26:52,760 --> 00:26:56,160 that allows the iron to be recycled 430 00:26:56,160 --> 00:26:59,240 to the different tissues. 431 00:26:59,240 --> 00:27:02,270 So we have a number of proteins that I'm 432 00:27:02,270 --> 00:27:05,840 going to very briefly introduce you to, 433 00:27:05,840 --> 00:27:08,270 in addition to these guys. 434 00:27:08,270 --> 00:27:14,030 And so we're getting into more acronyms cities. 435 00:27:14,030 --> 00:27:17,550 But the additional proteins that we need to think about-- 436 00:27:17,550 --> 00:27:33,400 so involved in iron homeostasis. 437 00:27:37,050 --> 00:27:42,530 Our number one, the ferritin, which 438 00:27:42,530 --> 00:27:44,948 in the introductory slide-- 439 00:27:44,948 --> 00:27:45,990 and let me just show you. 440 00:27:45,990 --> 00:27:47,350 So what I'm going to do, these are the list 441 00:27:47,350 --> 00:27:49,475 of proteins that I'm going to go through one by one 442 00:27:49,475 --> 00:27:51,230 and tell you a little bit. 443 00:27:51,230 --> 00:27:54,740 This is sort of an amazing protein. 444 00:27:54,740 --> 00:27:56,780 It has 24 protein subunits. 445 00:27:56,780 --> 00:27:58,640 It has two kinds of protein subunits. 446 00:27:58,640 --> 00:28:00,890 You don't need to remember this. 447 00:28:00,890 --> 00:28:03,110 But what is this function? 448 00:28:03,110 --> 00:28:06,440 It's a key-- and this is found in all organisms-- 449 00:28:06,440 --> 00:28:08,840 it's involved in iron storage. 450 00:28:11,920 --> 00:28:14,240 And why is this important? 451 00:28:14,240 --> 00:28:27,390 It's important because it keeps iron soluble so that it's not 452 00:28:27,390 --> 00:28:29,400 precipitating sort of as rust. 453 00:28:29,400 --> 00:28:32,550 There are, in yeast, if you look at some of yeast homeostasis, 454 00:28:32,550 --> 00:28:34,410 when things start going awry you can 455 00:28:34,410 --> 00:28:36,310 you can look at it in an electron microscope, 456 00:28:36,310 --> 00:28:38,930 you see iron all over the inside of the mitochondria, 457 00:28:38,930 --> 00:28:42,560 just these big black blobs where the iron has precipitated 458 00:28:42,560 --> 00:28:44,550 and mineralized. 459 00:28:44,550 --> 00:28:47,340 So we need to keep iron soluble, and we 460 00:28:47,340 --> 00:28:51,000 need to keep iron non-toxic. 461 00:28:51,000 --> 00:28:53,940 So what do I mean by non-toxic? 462 00:28:53,940 --> 00:28:57,150 In the last lecture, I told you that iron 2 can easily 463 00:28:57,150 --> 00:29:01,110 be oxidized to iron 3 by oxygen. We're going to talk about that 464 00:29:01,110 --> 00:29:03,570 in module 7 a little bit. 465 00:29:03,570 --> 00:29:07,260 And that can result in all kinds of damage inside the cell 466 00:29:07,260 --> 00:29:09,070 if it's not controlled. 467 00:29:09,070 --> 00:29:13,740 So this protein is sort of amazing. 468 00:29:13,740 --> 00:29:19,680 You can bind 4,500 irons, most of them 469 00:29:19,680 --> 00:29:23,310 are in the iron 3 state. 470 00:29:23,310 --> 00:29:26,850 But when you start out, it binds iron 2. 471 00:29:26,850 --> 00:29:29,340 So iron 2, again, inside the cell 472 00:29:29,340 --> 00:29:33,450 is what gets transferred around in general. 473 00:29:33,450 --> 00:29:40,260 So iron 2 binds, and then each ferritin 474 00:29:40,260 --> 00:29:42,390 has an oxidase activity that I'm not 475 00:29:42,390 --> 00:29:46,650 going to go into in detail that can oxidize it to iron 3, which 476 00:29:46,650 --> 00:29:48,690 puts it into this mineral structure 477 00:29:48,690 --> 00:29:52,250 that you see in these 4,500 atoms of iron. 478 00:29:52,250 --> 00:29:55,140 OK, you don't see it there, all you see is the protein there. 479 00:29:55,140 --> 00:30:01,580 So this gets oxidized to iron 3, and this is how 480 00:30:01,580 --> 00:30:07,810 it's stored in mineral form. 481 00:30:10,590 --> 00:30:13,770 So now the question is, say you needed iron. 482 00:30:13,770 --> 00:30:15,960 So we have a lot of iron, we want 483 00:30:15,960 --> 00:30:18,840 to keep it sequestered so we don't 484 00:30:18,840 --> 00:30:22,310 have to worry about reactive-- 485 00:30:22,310 --> 00:30:24,870 it doing chemistry that's aberrant. 486 00:30:24,870 --> 00:30:28,380 We want to keep it soluble. 487 00:30:28,380 --> 00:30:32,280 So we have iron stored in the plus 3 state 488 00:30:32,280 --> 00:30:33,780 in some kind of mineral form. 489 00:30:33,780 --> 00:30:35,680 How would you, if you wanted to use iron, 490 00:30:35,680 --> 00:30:36,720 now what would you do? 491 00:30:39,620 --> 00:30:42,430 Do you think you can get it out of the iron 3 mineral? 492 00:30:42,430 --> 00:30:43,010 No. 493 00:30:43,010 --> 00:30:46,710 What do you have to do to it to make the ligands more labile? 494 00:30:46,710 --> 00:30:48,530 All you need to reduce it. 495 00:30:48,530 --> 00:30:52,130 So to use it, you now-- and people are still 496 00:30:52,130 --> 00:30:54,110 arguing about what the reductants are-- 497 00:30:54,110 --> 00:31:09,100 so you need to reduce iron 2 plus 2 so you can use it. 498 00:31:17,780 --> 00:31:19,330 So that's ferritin. 499 00:31:19,330 --> 00:31:21,680 Does anybody have any questions about ferritin? 500 00:31:21,680 --> 00:31:24,770 It's got a complex structure, we have lots of structures of it. 501 00:31:24,770 --> 00:31:27,380 You can have-- every ferritin is sort of different, 502 00:31:27,380 --> 00:31:29,630 it has different ways of dealing with these issues 503 00:31:29,630 --> 00:31:32,880 of how you mineralize, and how you remove it. 504 00:31:32,880 --> 00:31:34,880 But this is a major storage protein 505 00:31:34,880 --> 00:31:37,460 in all organisms of ferritins. 506 00:31:37,460 --> 00:31:40,370 It's sort of an amazing structure. 507 00:31:40,370 --> 00:31:42,770 So what we were talking about before 508 00:31:42,770 --> 00:31:45,620 is that we get iron 3 transferrin. 509 00:31:45,620 --> 00:31:48,020 What does iron 3 transferrin look like? 510 00:31:48,020 --> 00:31:52,850 So we take iron from the diet, or we're recycling iron 511 00:31:52,850 --> 00:31:54,680 from red blood cells. 512 00:31:54,680 --> 00:31:57,530 We need to get it to the plus 3 state, where it gets picked up 513 00:31:57,530 --> 00:31:58,850 by transferrin. 514 00:31:58,850 --> 00:32:00,960 That's what we need to do. 515 00:32:00,960 --> 00:32:02,240 And so if you look at this-- 516 00:32:07,200 --> 00:32:13,900 So we've picked up iron 3 in transferrin. 517 00:32:17,330 --> 00:32:19,640 And the structures of transferrin are known. 518 00:32:19,640 --> 00:32:23,030 So now we need to look at transferrin-- 519 00:32:25,620 --> 00:32:28,310 whoops. 520 00:32:28,310 --> 00:32:30,350 And if you look at the structure, 521 00:32:30,350 --> 00:32:33,890 it is composed-- the protein is composed 522 00:32:33,890 --> 00:32:38,300 of two domains, each of which can bind iron 3 bicarbonate. 523 00:32:38,300 --> 00:32:42,170 So it has two little lobes over here. 524 00:32:42,170 --> 00:32:44,840 You can see this lobe and this lobe, the N terminal and the C 525 00:32:44,840 --> 00:32:46,040 terminal lobe. 526 00:32:46,040 --> 00:32:47,990 And they each bind-- 527 00:32:47,990 --> 00:32:50,870 if you look at this carefully, there is the iron, 528 00:32:50,870 --> 00:32:53,330 there is the bicarbonate. 529 00:32:53,330 --> 00:32:55,380 It has two tyrosines, a histidine, 530 00:32:55,380 --> 00:32:57,800 and an aspartate as ligands. 531 00:32:57,800 --> 00:33:01,040 And it's in an octahedral environment. 532 00:33:01,040 --> 00:33:04,220 So again, why bicarbonate? 533 00:33:04,220 --> 00:33:07,370 And people thought for a long time the bicarbonate was 534 00:33:07,370 --> 00:33:12,440 related potentially to how do you deliver this iron 3 535 00:33:12,440 --> 00:33:15,740 out of the transferrin into something that's useful, 536 00:33:15,740 --> 00:33:17,660 namely the enzymes that are going to use 537 00:33:17,660 --> 00:33:20,300 it to catalyze transformations. 538 00:33:20,300 --> 00:33:24,580 And what is the bi-- is there a role for bicarbonate 539 00:33:24,580 --> 00:33:26,380 in that process? 540 00:33:26,380 --> 00:33:28,510 So what's unusual about the transferrin, 541 00:33:28,510 --> 00:33:31,990 again, I get-- the KD is tight. 542 00:33:31,990 --> 00:33:37,210 What's most unusual is it's got bicarbonate, 543 00:33:37,210 --> 00:33:43,150 it's got two tyrosines, and it's got a histidine, 544 00:33:43,150 --> 00:33:50,625 and it's got an aspartate, and it's an octahedral environment. 545 00:33:50,625 --> 00:33:53,000 And how do you think-- what do you think the proteination 546 00:33:53,000 --> 00:33:55,250 state of the tyrosines are? 547 00:33:55,250 --> 00:33:57,985 Everybody know what tyrosine is? 548 00:33:57,985 --> 00:33:59,450 Do you think it's proteinated? 549 00:33:59,450 --> 00:34:00,290 Non-proteinated? 550 00:34:00,290 --> 00:34:03,080 This brings up another sort of general principle 551 00:34:03,080 --> 00:34:05,210 we talked about last time. 552 00:34:05,210 --> 00:34:08,270 If you have water attached to a metal, 553 00:34:08,270 --> 00:34:11,889 what can it do to the pKa of the water? 554 00:34:15,679 --> 00:34:18,800 It decreases it so that you lose the proton 555 00:34:18,800 --> 00:34:20,840 under physiological conditions. 556 00:34:20,840 --> 00:34:22,820 What's the pKa of tyronsine? 557 00:34:22,820 --> 00:34:25,969 It's on the order of 10, 10 1/2. 558 00:34:25,969 --> 00:34:29,960 And in fact, this is bound-- it's the phenylate. 559 00:34:29,960 --> 00:34:32,540 So both of these are in the phenylate form. 560 00:34:32,540 --> 00:34:35,210 So both of these are phenylate. 561 00:34:39,639 --> 00:34:41,780 And again, if you want to think more about this, 562 00:34:41,780 --> 00:34:44,630 both Liz and Lippert have taught a course, 563 00:34:44,630 --> 00:34:47,329 are teaching a course now, in bio inorganic chemistry, where 564 00:34:47,329 --> 00:34:50,750 you really sort of talk about the details of these kinds 565 00:34:50,750 --> 00:34:53,870 of interactions, which are key to the way everything 566 00:34:53,870 --> 00:34:55,489 functions. 567 00:34:55,489 --> 00:34:58,130 So we have transferrin, and the unusual part 568 00:34:58,130 --> 00:35:03,160 is the binding of bicarbonate, and then, again, 569 00:35:03,160 --> 00:35:06,950 let me just re-emphasize it's in the plus 3 state, 570 00:35:06,950 --> 00:35:09,800 and you have fairly tight binding. 571 00:35:09,800 --> 00:35:13,280 And what we're going to see is, it's going to bind just 572 00:35:13,280 --> 00:35:17,630 like the LDL particles bind to the LDL receptors, 573 00:35:17,630 --> 00:35:20,720 it's going to bind to the transferrin receptor. 574 00:35:20,720 --> 00:35:23,135 So we now have a transferrin receptor. 575 00:35:28,050 --> 00:35:29,050 So this is the receptor. 576 00:35:32,280 --> 00:35:36,810 And we know we have structures, actually, of the receptors. 577 00:35:36,810 --> 00:35:40,890 It's a 90 kilodalton dimer. 578 00:35:40,890 --> 00:35:42,470 So and its transmembrane. 579 00:35:42,470 --> 00:35:48,800 So you have-- so this is the transferrin receptor. 580 00:35:48,800 --> 00:35:52,900 I'm going to show you a cartoon of this in a minute. 581 00:35:52,900 --> 00:35:58,450 90 killodalton dimer, and so this is extracellular. 582 00:36:01,120 --> 00:36:02,740 This is intracellular. 583 00:36:05,678 --> 00:36:06,720 And this is the membrane. 584 00:36:12,890 --> 00:36:16,440 So let me just show you that cartoon over here. 585 00:36:16,440 --> 00:36:21,000 So extracellular, intracellular. 586 00:36:21,000 --> 00:36:24,750 And if you remember back to the LDL receptor, 587 00:36:24,750 --> 00:36:28,200 how did we trigger receptor mediated endocytosis? 588 00:36:28,200 --> 00:36:30,330 We had a zip code. 589 00:36:30,330 --> 00:36:32,460 Here we also have a zip code. 590 00:36:32,460 --> 00:36:35,790 And the zip code is YTRF. 591 00:36:35,790 --> 00:36:41,000 So there's also, on the intracellular side, 592 00:36:41,000 --> 00:36:54,490 a zip code for triggering transferrin uptake. 593 00:36:59,890 --> 00:37:04,210 So those are the players that we need to think about. 594 00:37:04,210 --> 00:37:06,460 So the transferrin, in the transferrin receptor, 595 00:37:06,460 --> 00:37:08,200 have parallels with LDL. 596 00:37:08,200 --> 00:37:10,480 LDL receptor-- of course every one of these things 597 00:37:10,480 --> 00:37:12,280 is different. 598 00:37:12,280 --> 00:37:15,100 But this was one of the other systems that 599 00:37:15,100 --> 00:37:17,740 had been characterized quite extensively, 600 00:37:17,740 --> 00:37:20,900 the first one being the LDL receptor. 601 00:37:20,900 --> 00:37:24,650 And so the model is shown here. 602 00:37:24,650 --> 00:37:27,580 This model hasn't really been-- 603 00:37:27,580 --> 00:37:29,272 this model's not completely correct. 604 00:37:29,272 --> 00:37:31,730 I'll tell you where things need to be changed a little bit. 605 00:37:31,730 --> 00:37:33,550 But really people haven't studied this model 606 00:37:33,550 --> 00:37:35,008 in a long time, even though there's 607 00:37:35,008 --> 00:37:36,550 a lot we don't understand. 608 00:37:36,550 --> 00:37:38,290 So here's the surface. 609 00:37:38,290 --> 00:37:40,960 Here's transferrrin, these little things here. 610 00:37:40,960 --> 00:37:44,340 Here's the transferrin receptor purple. 611 00:37:44,340 --> 00:37:48,990 So the transferrin binds to the transferrin receptor. 612 00:37:48,990 --> 00:37:53,670 To get uptake into the cell, you need to have clustering. 613 00:37:53,670 --> 00:37:56,340 So that's not shown here, because this cartoon 614 00:37:56,340 --> 00:37:59,770 was drawn before we realized that you had a cluster-- 615 00:37:59,770 --> 00:38:02,460 the transferrin receptors. 616 00:38:02,460 --> 00:38:05,610 When you transfer, when you cluster, 617 00:38:05,610 --> 00:38:09,870 and you bind transferrin, again, just like we saw with the LDL 618 00:38:09,870 --> 00:38:12,750 receptor, in some way, you have machinery 619 00:38:12,750 --> 00:38:16,890 that attracts the clathrin, and then it's 620 00:38:16,890 --> 00:38:21,120 going to pinch off the clathrin coated vesicle. 621 00:38:21,120 --> 00:38:25,710 And they skip here the clathrin coated vesicle. 622 00:38:25,710 --> 00:38:27,750 So that should be in between-- this is clathrin. 623 00:38:30,270 --> 00:38:33,330 And then what happens, just like in the LDL receptor, 624 00:38:33,330 --> 00:38:36,840 you remove the clathrin from the external part 625 00:38:36,840 --> 00:38:38,540 of your little vesicle. 626 00:38:38,540 --> 00:38:40,710 So that's what's indicated here. 627 00:38:40,710 --> 00:38:41,860 So what do we have? 628 00:38:41,860 --> 00:38:47,550 We have the transferrin receptor, and transferrin, 629 00:38:47,550 --> 00:38:53,430 and this is-- the internal pH of this system is about 5-5. 630 00:38:53,430 --> 00:38:57,990 So if you think about this, how would you-- 631 00:38:57,990 --> 00:39:01,140 how would you remove the iron from the transferrin? 632 00:39:01,140 --> 00:39:02,910 Why might bicarbonate be there? 633 00:39:02,910 --> 00:39:06,315 So I just told you that bicarbonate in iron 634 00:39:06,315 --> 00:39:08,520 are bound to the transferrin. 635 00:39:08,520 --> 00:39:11,700 Can you think of a mechanism by which that could happen? 636 00:39:11,700 --> 00:39:16,090 X inside the cell, at lower pH? 637 00:39:16,090 --> 00:39:17,560 We don't know the answer to this. 638 00:39:17,560 --> 00:39:19,450 It's still open to debate. 639 00:39:19,450 --> 00:39:23,470 So-- but what happens to the bicarbonate at low pH? 640 00:39:27,850 --> 00:39:30,490 Think about hemoglobin. 641 00:39:30,490 --> 00:39:32,680 Think about 5.07 and hemoglobin. 642 00:39:32,680 --> 00:39:37,390 We spend so much time talking about bicarbonate 643 00:39:37,390 --> 00:39:41,800 as a key player inside red blood cells. 644 00:39:41,800 --> 00:39:44,140 What happens to bicarbonate in the presence of acid? 645 00:39:51,160 --> 00:39:53,170 Yeah, so it forms carbonic acid. 646 00:39:53,170 --> 00:39:55,652 What happened to the carbonic acid? 647 00:39:55,652 --> 00:39:57,310 To CO2 in the water. 648 00:39:57,310 --> 00:39:57,820 Yeah. 649 00:39:57,820 --> 00:39:59,680 So this is something we banged into you 650 00:39:59,680 --> 00:40:01,890 over and over again in 5.07. 651 00:40:01,890 --> 00:40:05,800 There's an equilibrium that happens over and over again 652 00:40:05,800 --> 00:40:06,430 inside cells. 653 00:40:06,430 --> 00:40:08,980 So maybe that's a way to deliver the iron. 654 00:40:08,980 --> 00:40:10,750 I don't know. 655 00:40:10,750 --> 00:40:14,560 So we somehow lose iron. 656 00:40:14,560 --> 00:40:18,330 But the iron is in the plus 3 state. 657 00:40:18,330 --> 00:40:21,750 To get it into the cytosol, which 658 00:40:21,750 --> 00:40:23,880 is where we're going to use it, to deliver it 659 00:40:23,880 --> 00:40:26,880 to all of the proteins, what do we need to do? 660 00:40:26,880 --> 00:40:28,980 Hopefully you now remember this. 661 00:40:28,980 --> 00:40:31,200 We need to reduce it. 662 00:40:31,200 --> 00:40:38,790 So steep is a reductase, a ferric reductase, 663 00:40:38,790 --> 00:40:41,040 that converts this into iron 2. 664 00:40:41,040 --> 00:40:43,040 Where did we see this guy before? 665 00:40:43,040 --> 00:40:44,950 DMT1. 666 00:40:44,950 --> 00:40:48,810 We've see that before as a key player in uptake 667 00:40:48,810 --> 00:40:49,740 into enterocytes. 668 00:40:49,740 --> 00:40:53,580 So you see these same players over and over again. 669 00:40:53,580 --> 00:40:57,630 You see this shift from iron 2 to iron 3 over and over again, 670 00:40:57,630 --> 00:41:01,230 actually, in yeast, where I know a lot about iron metabolism 671 00:41:01,230 --> 00:41:04,310 as well as in human systems. 672 00:41:04,310 --> 00:41:11,130 Now-- so we've got iron 2 out of the transferrin, transferrin 673 00:41:11,130 --> 00:41:12,150 receptor. 674 00:41:12,150 --> 00:41:15,450 And then the iron 2 goes into the cytosol. 675 00:41:15,450 --> 00:41:18,090 And then we've got to figure out how to use it in a way 676 00:41:18,090 --> 00:41:19,980 so that we don't have oxidative stress 677 00:41:19,980 --> 00:41:22,380 and deliver it to the proteins to biosynthesize 678 00:41:22,380 --> 00:41:25,350 all our co-factors. 679 00:41:25,350 --> 00:41:28,125 So then the question is, remember in the LDL receptor, 680 00:41:28,125 --> 00:41:30,760 it got recycled. 681 00:41:30,760 --> 00:41:33,190 So what happens here is distinct from what 682 00:41:33,190 --> 00:41:36,070 happens in the LDL receptor. 683 00:41:36,070 --> 00:41:42,990 In that now the transferrin and the transferrin receptor 684 00:41:42,990 --> 00:41:45,750 are both recycled. 685 00:41:45,750 --> 00:41:48,900 So that's distinct from what we briefly 686 00:41:48,900 --> 00:41:52,220 talked about in the case of cholesterol. 687 00:41:52,220 --> 00:41:57,450 So we have two ways of taking iron into the cell one-- 688 00:41:57,450 --> 00:41:59,810 is through these di-- 689 00:41:59,810 --> 00:42:04,260 iron 2 transporters, the DMT molecules, and the second way 690 00:42:04,260 --> 00:42:07,230 is through iron transfer-- iron transferrin 691 00:42:07,230 --> 00:42:09,840 which circulates in the blood and delivers it 692 00:42:09,840 --> 00:42:11,730 to all the tissues. 693 00:42:11,730 --> 00:42:15,540 So these are the major mechanisms of iron delivery, 694 00:42:15,540 --> 00:42:20,070 and recycling within the cell controlled by hepcidin, 695 00:42:20,070 --> 00:42:24,370 this peptide hormone. 696 00:42:24,370 --> 00:42:29,220 So what I want to do now is look at how this iron is sensed. 697 00:42:29,220 --> 00:42:31,520 How do we control everything? 698 00:42:31,520 --> 00:42:32,570 And iron sensing-- 699 00:42:36,935 --> 00:42:39,060 So iron sensing, there are going to be two players. 700 00:42:41,708 --> 00:42:43,500 And so we're going to look at iron sensing. 701 00:42:49,050 --> 00:42:51,610 And I'm going to introduce you to the two players, 702 00:42:51,610 --> 00:42:55,210 and then I'm going to show you the general logic of how you 703 00:42:55,210 --> 00:43:00,850 control all these proteins we've talked about-- ferritin, DMT1, 704 00:43:00,850 --> 00:43:02,260 transferrin receptor-- 705 00:43:02,260 --> 00:43:04,780 all of these things are going to be controlled 706 00:43:04,780 --> 00:43:06,400 by the mechanism we're going to talk 707 00:43:06,400 --> 00:43:11,290 about now, which is regulation at the translational level. 708 00:43:11,290 --> 00:43:21,610 So this is iron sensing by translational control. 709 00:43:21,610 --> 00:43:24,610 So who are the two players? 710 00:43:24,610 --> 00:43:26,750 They're written up there. 711 00:43:26,750 --> 00:43:30,260 But we have iron responsive element, 712 00:43:30,260 --> 00:43:33,440 and we're going to see that's a little piece of RNA. 713 00:43:33,440 --> 00:43:35,550 So-- and I'll show you what it looks like. 714 00:43:35,550 --> 00:43:39,200 So this is RNA, a little piece of RNA, 715 00:43:39,200 --> 00:43:42,650 stem loop piece of RNA, that has defined characteristics. 716 00:43:42,650 --> 00:43:45,050 I'm going to show you what it is. 717 00:43:45,050 --> 00:43:51,110 And then we have iron responsive protein 1, 718 00:43:51,110 --> 00:43:55,270 or iron responsive binding protein 1. 719 00:43:55,270 --> 00:43:56,770 They're called both of these things, 720 00:43:56,770 --> 00:43:59,710 I don't remember what was in the articles you had to read. 721 00:43:59,710 --> 00:44:02,180 They're sort of used interchangeably. 722 00:44:02,180 --> 00:44:06,307 And there are two of these, so there's a 1 and there's a 2. 723 00:44:06,307 --> 00:44:08,390 And they're structurally homologous to each other, 724 00:44:08,390 --> 00:44:10,880 and I'll tell you a little bit about each one of these. 725 00:44:10,880 --> 00:44:15,887 So we also have a one and a two. 726 00:44:15,887 --> 00:44:16,970 So those are the two guys. 727 00:44:16,970 --> 00:44:18,410 These are proteins. 728 00:44:18,410 --> 00:44:22,160 So these are proteins, that's why the name binding protein. 729 00:44:22,160 --> 00:44:31,610 So it turns out that iron responsive binding proteins 730 00:44:31,610 --> 00:44:44,180 are homologous to aconitase-- 731 00:44:44,180 --> 00:44:47,330 where you seen aconitase before? 732 00:44:47,330 --> 00:44:49,660 Yeah, so in the TCA cycle. 733 00:44:49,660 --> 00:44:55,798 It catalyzes the conversion of citrate to isocitrate. 734 00:44:55,798 --> 00:44:59,370 So-- and where is the TCA-- 735 00:44:59,370 --> 00:45:00,840 TCA cycle located? 736 00:45:00,840 --> 00:45:02,650 In the mitochondria. 737 00:45:02,650 --> 00:45:13,860 So this is a TCA cycle enzyme found in the mitochondria. 738 00:45:17,090 --> 00:45:21,470 But what we'll see is, we're working on RNA, 739 00:45:21,470 --> 00:45:23,680 we're going to regulate somehow. 740 00:45:23,680 --> 00:45:26,600 We're going to use interaction between this protein 741 00:45:26,600 --> 00:45:29,660 and a piece of RNA to control the translational process, 742 00:45:29,660 --> 00:45:32,510 where is that located in the cytosol? 743 00:45:32,510 --> 00:45:36,180 So these proteins are located in the cytosol. 744 00:45:40,010 --> 00:45:46,060 So if you think about what happens with aconitase, 745 00:45:46,060 --> 00:45:49,333 let me just write that down for you. 746 00:45:49,333 --> 00:45:50,125 So we have citrate. 747 00:45:54,859 --> 00:45:58,810 And I asked the question, do you think 748 00:45:58,810 --> 00:46:04,720 it's interesting that citrate is involved 749 00:46:04,720 --> 00:46:07,120 in this overall process that I'm going to be describing? 750 00:46:07,120 --> 00:46:09,100 What do we know about citrate, besides the fact 751 00:46:09,100 --> 00:46:11,845 that it's an intermediate in the TCA cycle? 752 00:46:15,100 --> 00:46:17,640 So this is citrate. 753 00:46:17,640 --> 00:46:20,320 It undergoes a dehydration reaction. 754 00:46:20,320 --> 00:46:29,050 So we're going to lose water to form aconitate, cis-aconitate-- 755 00:46:36,870 --> 00:46:38,760 and then it becomes rehydrated. 756 00:46:42,290 --> 00:46:49,460 So that's the reaction you learned about a long time ago 757 00:46:49,460 --> 00:46:53,260 in the Krebs cycle or the TCA cycle. 758 00:46:53,260 --> 00:46:56,057 Why is it interesting that citrate is involved? 759 00:46:56,057 --> 00:46:57,640 I don't know why it's really involved. 760 00:46:57,640 --> 00:46:59,110 But do you think it's interesting? 761 00:46:59,110 --> 00:47:03,020 What is citrate, if you look at the structure of it? 762 00:47:03,020 --> 00:47:03,520 Yeah. 763 00:47:03,520 --> 00:47:04,310 AUDIENCE: Combined iron. 764 00:47:04,310 --> 00:47:05,768 JOANNE STUBBE: Yeah, combined iron. 765 00:47:05,768 --> 00:47:08,320 And in fact, there are iron siderophores that use citrate. 766 00:47:08,320 --> 00:47:10,120 I don't think this is an accident. 767 00:47:10,120 --> 00:47:13,240 And thinking about, again, how nature uses primary 768 00:47:13,240 --> 00:47:16,510 metabolites over and over again in ways other than what 769 00:47:16,510 --> 00:47:20,200 you see in primary metabolism. 770 00:47:20,200 --> 00:47:24,590 So what's unusual about this protein is the following. 771 00:47:24,590 --> 00:47:27,670 And this is the key to the way the sensing is 772 00:47:27,670 --> 00:47:31,510 going to work for the iron responsive binding proteins. 773 00:47:31,510 --> 00:47:33,820 So if you look at-- if you go back and you look-- 774 00:47:36,690 --> 00:47:39,698 if you go back and you look at the Krebs cycle, or you go back 775 00:47:39,698 --> 00:47:41,490 and you think about this, this is something 776 00:47:41,490 --> 00:47:43,980 that probably confused you all. 777 00:47:49,960 --> 00:47:55,720 You have an iron 3, a 4 iron 4 sulfur cluster. 778 00:47:55,720 --> 00:47:58,270 Remember I talked a little bit about this, 779 00:47:58,270 --> 00:48:00,040 trying to show you that this was going 780 00:48:00,040 --> 00:48:05,270 to be highlighted later on? 781 00:48:08,120 --> 00:48:11,300 and what we have in this 4 iron 4 sulfur cluster-- 782 00:48:17,950 --> 00:48:22,360 you have a cysteine attached to three of the irons. 783 00:48:22,360 --> 00:48:26,070 We have one iron that's unique, OK 784 00:48:26,070 --> 00:48:30,270 that doesn't have the cysteine that you see in normal 4 iron 4 785 00:48:30,270 --> 00:48:31,170 sulfur clusters. 786 00:48:31,170 --> 00:48:32,790 So this is the unique iron. 787 00:48:38,870 --> 00:48:40,850 So if you look at that over here-- so here's 788 00:48:40,850 --> 00:48:43,010 the cartoon of this. 789 00:48:43,010 --> 00:48:45,920 So here you have cysteine, cysteine, cysteine in the 4 790 00:48:45,920 --> 00:48:47,710 iron 4 sulfur cluster. 791 00:48:47,710 --> 00:48:49,810 Here's citrate. 792 00:48:49,810 --> 00:48:51,850 And that iron-- so most of you probably 793 00:48:51,850 --> 00:48:54,220 learned in respiration, iron sulfur clusters 794 00:48:54,220 --> 00:48:56,350 are involved in electron transfer. 795 00:48:56,350 --> 00:48:57,880 They do one electron chemistry. 796 00:48:57,880 --> 00:49:01,060 They undergo oxidation reduction, which we briefly 797 00:49:01,060 --> 00:49:04,100 discussed in the last lecture. 798 00:49:04,100 --> 00:49:06,040 But what's it doing here? 799 00:49:06,040 --> 00:49:09,760 What it's doing here is binding the citrate. 800 00:49:09,760 --> 00:49:11,410 So here's citrate. 801 00:49:11,410 --> 00:49:14,350 This is the hydroxyl that we're going 802 00:49:14,350 --> 00:49:20,510 to eliminate to lose water to form cis-aconitate. 803 00:49:20,510 --> 00:49:23,120 So this is the first example. 804 00:49:23,120 --> 00:49:25,730 But this was discovered by Helmut Beinert at Wisconsin 805 00:49:25,730 --> 00:49:28,540 many years ago, where the iron sulfur 806 00:49:28,540 --> 00:49:32,480 classes were doing something other than redox chemistry. 807 00:49:32,480 --> 00:49:33,980 This is just the tip of the iceberg. 808 00:49:33,980 --> 00:49:37,410 Remember, I talked to you about radical SAM proteins, 809 00:49:37,410 --> 00:49:39,920 100,000 proteins doing interesting chemistry. 810 00:49:39,920 --> 00:49:41,750 This is the first example of this. 811 00:49:41,750 --> 00:49:44,330 And these really are seminal experiments 812 00:49:44,330 --> 00:49:46,850 to figure out how this all worked. 813 00:49:46,850 --> 00:49:51,980 So the unusual thing is that most iron sulfur clusters look 814 00:49:51,980 --> 00:49:55,280 like this, and they all have 16 on each of the iron, 815 00:49:55,280 --> 00:49:57,650 and they do redox chemistry, but now we're 816 00:49:57,650 --> 00:50:01,300 finding that a lot of iron sulfur clusters 817 00:50:01,300 --> 00:50:04,460 have unique iron they can end up doing interesting chemistry as 818 00:50:04,460 --> 00:50:08,750 well, namely binding S-adenosyl methionine. 819 00:50:08,750 --> 00:50:12,140 So if you go back and you think about what happens, 820 00:50:12,140 --> 00:50:14,830 this is helping dehydration. 821 00:50:14,830 --> 00:50:17,740 So you're going to dehydrate. 822 00:50:17,740 --> 00:50:19,690 But now you have to reorganize the thing. 823 00:50:19,690 --> 00:50:22,630 This is one where they talk about the Ferris-- 824 00:50:22,630 --> 00:50:24,670 spinning around the Ferris wheel if you 825 00:50:24,670 --> 00:50:27,220 look at an introductory TCA cycle thing, 826 00:50:27,220 --> 00:50:28,600 how this reorganizes. 827 00:50:28,600 --> 00:50:30,372 I don't think this is a very good picture. 828 00:50:30,372 --> 00:50:32,080 But it needs to reorganize because you're 829 00:50:32,080 --> 00:50:36,280 going to rehydrate another carbon, using the same iron. 830 00:50:36,280 --> 00:50:40,240 So if you sit here and you stare at this, what you see 831 00:50:40,240 --> 00:50:42,460 is this carboxylate. 832 00:50:42,460 --> 00:50:45,490 Now, here was the initial carboxylate bound, 833 00:50:45,490 --> 00:50:47,140 this one wasn't bound. 834 00:50:47,140 --> 00:50:50,320 Now, this one ends up being bound. 835 00:50:50,320 --> 00:50:54,640 And now you're adding water back across this double bond. 836 00:50:54,640 --> 00:50:57,130 So the purpose of this system is simply 837 00:50:57,130 --> 00:51:00,310 to catalyze the dehydration reaction. 838 00:51:00,310 --> 00:51:03,790 So what the heck are we doing with an iron 839 00:51:03,790 --> 00:51:10,060 responsive binding protein being a cytosolic aconitase 840 00:51:10,060 --> 00:51:11,628 equivalent? 841 00:51:11,628 --> 00:51:13,420 And so what I'm going to come back and tell 842 00:51:13,420 --> 00:51:17,740 you one Friday is, this is going to be the key switch for iron 843 00:51:17,740 --> 00:51:18,670 sensing. 844 00:51:18,670 --> 00:51:22,660 Whether the iron is in the apostate, with no metal, 845 00:51:22,660 --> 00:51:27,310 or whether it moves to the 4 iron 4 sulfur cluster state. 846 00:51:27,310 --> 00:51:30,070 And we'll talk a little bit then about how 847 00:51:30,070 --> 00:51:34,360 those two states, and the presence of RNA, 848 00:51:34,360 --> 00:51:36,640 can control which of all these proteins 849 00:51:36,640 --> 00:51:39,940 I've thrown at you today actually get translated. 850 00:51:39,940 --> 00:51:41,490 OK.