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| 02:22.15 | omniter | hey guys. anyone had a problem entering their postal code for the prepaid card? |
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| 02:26.52 | ojwb | omniter: apparently it's google's postcode you should use |
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| 02:28.26 | Upthorn | ojwb: only if he lives outside the US |
| 02:28.39 | Upthorn | (s)he, I mean |
| 02:30.53 | ojwb | ah, ok |
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| 05:44.03 | kblin | yay |
| 05:44.17 | kblin | done with the initial draft of my thesis.. |
| 05:44.47 | kblin | time to print it, read it, freak out and start to rewrite it in a better way |
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| 05:55.13 | Mkop | kblin: thesis on what? |
| 05:55.46 | kblin | my current title is "Substrate specifity and RNA binding activity of Streptomyces virido\-chromogenes aconitases AcnA and PMI" |
| 05:56.40 | ojwb | will wait for the film |
| 05:57.11 | ojwb | but congratulations |
| 05:57.19 | Mkop | what's an aconitase? |
| 05:57.54 | kblin | it's an enzyme commonly found in the tricarboxylic acid cycle (citrate cycle) |
| 05:58.09 | kblin | it catalyzes the reaction of citrate to isocitrate |
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| 05:58.38 | kblin | in my streptomycete, the aconitase doing that is AcnA |
| 05:58.40 | Mkop | citrate is a C3? |
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| 05:59.32 | kblin | no, it's got a C3 backbone with three carboxy-groups |
| 05:59.34 | Mkop | in my bio class, I didn't have to know the names of all the different steps in the krebs/citric acid cycles, just C6 -> 2*C3 -> ... |
| 05:59.49 | Mkop | carboxy = COOH, right? |
| 05:59.54 | kblin | yeah |
| 06:00.14 | kblin | http://en.wikipedia.org/wiki/File:Citric_acid_cycle_with_aconitate_2.svg |
| 06:00.37 | kblin | aconitase does the citrate<->aconitate<->d-isocitrate step |
| 06:00.40 | Mkop | so what does RNA binding have to do with anything? |
| 06:00.49 | Mkop | I mean here particularly |
| 06:00.52 | thebolt | morning kblin |
| 06:01.33 | kblin | Mkop: well, a central feature of aconitases is a central 4Fe 4S cluster |
| 06:02.07 | kblin | Mkop: and one of the iron ions can easily leave the cluster, completely changing what aconitase is doing in a cell |
| 06:02.30 | kblin | at least for eukaryotes this has been shown for aconitase not in the mitochondria |
| 06:03.46 | Mkop | I remember something about an Fe cluster in whatever the last redox enzyme is in mitochondria |
| 06:03.48 | kblin | once the cluster is deactivated, the aconitase opens along a hinge region and starts binding to mRNA sequences found on genes related to cellular iron regulation |
| 06:03.56 | Mkop | enzyme IV, something like that? |
| 06:04.10 | Mkop | I've forgotten a lot, it seems |
| 06:04.29 | kblin | dunno really. biologists have like zillions of different names for the same things all over the place |
| 06:04.45 | Mkop | complex IV |
| 06:04.56 | Mkop | aka cytochrome C oxidase |
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| 06:05.03 | Mkop | what are you? chemist? |
| 06:05.14 | Mkop | no, that wouldn't make sense |
| 06:05.26 | Mkop | what you're doing is molecular biology, seems like.... |
| 06:06.12 | Mkop | what school? |
| 06:06.18 | Mkop | and I assume this is a PhD thesis? |
| 06:06.24 | kblin | in any case, as aconitase loses one Fe-ion when the iron concentration in the cell is low, aconitase suddenly starts to block the translation of mRNA related to storing iron and upregulates translation of mRNA related to proteins transporting iron into a cell |
| 06:06.51 | Mkop | cool! |
| 06:06.55 | kblin | I'm a computational biologist, University of Tübingen, Germany, and it's a M.Sc. thesis |
| 06:07.19 | kblin | yeah, biology is full of little miracles at cellular level |
| 06:07.44 | kblin | so anyway, this is for eukaryotes |
| 06:08.09 | Mkop | so when you say RNA binding, you mean the protein binding to the mRNA? |
| 06:08.20 | kblin | similiar RNA binding functionality has been shown for some bacterial aconitases |
| 06:08.50 | kblin | well, aconitase seems to bind to all RNA that folds into a specific 3D structure |
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| 06:09.15 | kblin | but usually the only RNA floating around in a cell that can fold to this structure is mRNA |
| 06:10.01 | Mkop | I went to this seminar about six months ago by computational biology guy who was looking at actin and microtubule stability from a quantum level |
| 06:10.34 | kblin | he, yeah, for 7 ns or the like |
| 06:10.37 | Mkop | starting from the stability and the amount of freedom allowed by different kinds of chemical bonds, etc. |
| 06:10.40 | kblin | unless he really had a lot of time |
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| 06:11.07 | kblin | molecular dynamics simulations are really CPU-heavy |
| 06:11.14 | Mkop | the force exerted by van der Waals forces vs. covalent vs. hydrogen bonds vs. sulfide bridges |
| 06:11.24 | Mkop | yeah, that's what I gathered |
| 06:11.44 | kblin | yeah, but you commonly cheat by using a force field to give approximate values for this |
| 06:12.16 | kblin | it gives ok results and is faster than real calculation by many orders of magnitude |
| 06:13.14 | Mkop | b/c actin/MTs are such ginormous things, (polymers of thousands of monomers long), they first did calculations to determine how much one monomer can move relative to another, and then used like 6 values (rotation, translation in each direction, times 2 or 3 monomers) to figure out what's gonna happen for the whole polymer |
| 06:13.35 | kblin | like if you wanted to do a full quantum-based simulation of pretty much anything that happens in a molecule the size of an enzyme, the sun would go out before you're finished |
| 06:14.48 | kblin | so as much as I'd hate somebody proving that P=NP from a cryptography point of view, as a computational biologist I'd be very happy about this |
| 06:15.18 | Mkop | P=NP> |
| 06:15.19 | Mkop | ? |
| 06:15.42 | dberkholz | so if there isn't enough iron around, aconitase loses its iron and tells the cell to stop making crap that needs iron? |
| 06:15.55 | kblin | dberkholz: yeah, pretty much |
| 06:16.01 | Mkop | tells it to stop storing iron |
| 06:16.23 | dberkholz | that's an interesting bifunctionality |
| 06:16.47 | kblin | dberkholz: and as soon as there's enough iron around, it switches back to it's normal function |
| 06:17.04 | kblin | you wouldn't want too much free iron in a cell either |
| 06:17.22 | dberkholz | indeed. |
| 06:17.37 | dberkholz | knows a bit about that sort of thing |
| 06:18.03 | kblin | Mkop: there's a long standing question if problems solvable with non-polinomial algorithms are solvable by polinomial algorithms or not |
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| 06:18.25 | kblin | Mkop: so far, no one has been able to prove or disprove it |
| 06:18.30 | dberkholz | i'm a fan of multiscale simulations. |
| 06:18.48 | Mkop | it's the same kind of regulation that happens with glucose. When there's a lot of glucose in the cell, the extra glucose activates promoters that tell the cell to activate genes that break down glucose, and vice versa |
| 06:19.05 | Mkop | kblin: now you're starting with the computer science stuff. :-( |
| 06:19.15 | Mkop | I'm a programmer and engineer, not a computer scientist :-P |
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| 06:19.31 | kblin | in any case, some bacterial aconitases seem to be able to bind to similar RNA structures |
| 06:20.06 | dberkholz | is the rna site masked during "normal" activity and vice versa? |
| 06:20.10 | Mkop | do they have the same genes for iron storage? |
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| 06:20.58 | kblin | dberkholz: it's pretty much at the same site, but it seems like once the Fe-S cluster is complete, RNA won't bind |
| 06:21.37 | kblin | and as soon as the Fe-S cluster is destroyed, the citrate activity is gone and RNA will bind |
| 06:21.43 | Mkop | thus, "substrate specificity" |
| 06:21.50 | kblin | no |
| 06:21.54 | dberkholz | kblin: is a complex structure known? |
| 06:22.01 | Mkop | ok, then no |
| 06:22.16 | Mkop | what's the substrate specificity referring to then? |
| 06:22.21 | dberkholz | my postdoc might be vaguely in the area of iron-sulfur clusters |
| 06:22.32 | kblin | dberkholz: yeah. 7ACN or 1C96 for citrate and 2IPY for RNA |
| 06:22.34 | Mkop | wikipedia has a picture of the protein |
| 06:23.09 | kblin | Mkop: my bacterium Streptomyces viridochromogenes has another aconitase-like protein |
| 06:23.14 | kblin | that's the PMI |
| 06:23.48 | kblin | it's got a 55% sequence identity on the protein level, which is like "pretty much the same thing" in protein scale |
| 06:24.05 | thebolt | hehe |
| 06:24.20 | Mkop | 55% sequence identity = 55% of the amino acids match up? |
| 06:24.25 | kblin | yeah |
| 06:24.35 | thebolt | wonder if my teacher in say digital communication would accept that "oh well, the received signal is 55% identic to the sent signal.. pretty much the same thing" ;) |
| 06:25.36 | kblin | thebolt: if you compare cytosolic aconitases to mitochondrial aconitases, they actuall do the same thing and are only 27% identical on the sequence level |
| 06:25.53 | kblin | well, mitochondral aconitases don't do the RNA binding part |
| 06:26.03 | kblin | but the citric acid cycle activity is the same |
| 06:26.16 | kblin | and the structures match up pretty well |
| 06:26.45 | kblin | so I've got a pretty decent level of confidence in my 55% identical protein looking the same |
| 06:27.01 | thebolt | kblin: :) |
| 06:27.19 | thebolt | and we want <10^-3 error rates (or better..) |
| 06:27.26 | Mkop | the question is, do the important regions exhibit very high sequence identity? |
| 06:27.34 | kblin | especially as my biologists could show that the RNA binding activity on both S. viridochromogenes aconitases are the same |
| 06:28.01 | kblin | Mkop: the active site has 18 of 20 conserved AA being identical |
| 06:28.13 | Mkop | when you quantify sequence identity with a number like that, do you take into account similar amino acids? |
| 06:28.43 | Mkop | AA? |
| 06:28.43 | kblin | no, if I look at conservative mutations, identity is around 70% |
| 06:28.49 | kblin | amino acids |
| 06:28.56 | Mkop | oh, of course |
| 06:29.07 | dberkholz | hm, yeah, that is fairly close. |
| 06:29.08 | kblin | sorry, it's easy to get carried away |
| 06:29.22 | dberkholz | citrate sure looks tiny next to that humongous rna |
| 06:29.42 | kblin | now the fun part is that my other aconitase doesn't like citrate at all |
| 06:29.57 | Mkop | "conservative mutations" means taking into account that one amino acid can easily mutate into another of the same class with very little effect? |
| 06:30.17 | kblin | it instead catalyzes the reaction of phosphinometyl malate to isophosphinometyl malate |
| 06:30.19 | r0bby | AWESOMEEEEEEEEEEEEEEEEEEEEEEEEE |
| 06:30.20 | kblin | Mkop: yeah |
| 06:30.20 | Mkop | dberkholz: what picture are you looking at? |
| 06:30.23 | r0bby | time to fail :D |
| 06:30.45 | kblin | and that's what the "substrate specifity" part is about |
| 06:30.52 | dberkholz | Mkop: i opened up the two protein structures in a pdb viewer |
| 06:31.01 | Mkop | oh |
| 06:31.12 | Mkop | hmm, I wonder if I have access to a pdb viewer |
| 06:31.18 | Mkop | probably could if I cared to |
| 06:31.33 | dberkholz | look around for jmol, there should be a java applet somewhere |
| 06:31.37 | kblin | Mkop: if you've got a java plugin, PDB has a java applet |
| 06:32.22 | dberkholz | i don't think overlaying the structures is very straightforward though |
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| 06:32.53 | kblin | now phosphinomethyl malate looks pretty much the same as citrate, just that one carboxy-group is replaced by a phosphino-group |
| 06:33.28 | Mkop | what's phosphino? |
| 06:33.49 | kblin | dberkholz: I've used a multiple sequence alignment of multiple aconitases to the sequence of PDB structure 2B3Y to identify active site residues |
| 06:33.59 | kblin | Mkop: HOPOH |
| 06:34.32 | Mkop | P has 3 free bonds, right? |
| 06:34.47 | kblin | well, depends :) |
| 06:35.01 | Mkop | phosphate is PO4 |
| 06:35.21 | kblin | in this case the P is bound to the malate-backbone |
| 06:35.32 | kblin | so there's one free electron pair |
| 06:35.47 | Mkop | it's bound to the backbone, or it's part of the backbone? |
| 06:36.12 | kblin | it's bound to the backbone instead of one COOH |
| 06:36.40 | kblin | now, a phoshino group has almost the same size as a carboxy group |
| 06:37.25 | kblin | given this and the fact that docking programs seem to choke on the [Fe-S] cluster alot, you can bet I had a lot of fun with this |
| 06:37.58 | kblin | in fact I'm still waiting for experimental results on my proposed mutations to prove substrate specifity |
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| 06:38.16 | kblin | because if the mutations I proposed don't work out, I don't have any bloody clue :) |
| 06:38.49 | Mkop | kblin: how long is your thesis? |
| 06:39.40 | thebolt | kblin: so then you are smoked? ;-) (or have to rewrite your report to show you are wrong ;) |
| 06:40.00 | kblin | Mkop: 38 pages, with pictures and all |
| 06:40.16 | Mkop | oh, that's not so long |
| 06:41.11 | Mkop | considering PhD theses can easily be 300 |
| 06:41.11 | kblin | thebolt: the docking results didn't tell anything. so if my proposed mutations don't work, I can always claim this was already visible in the docking results ;) |
| 06:41.11 | dberkholz | Mkop: if you install pymol (most distros have it), i can send you a session file to open that'll let you look at this. |
| 06:41.33 | kblin | thebolt: if the mutations work, I can claim that docking programs can't deal with the Fe-S cluster and the substrates |
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| 06:43.10 | Mkop | lol |
| 06:43.36 | Mkop | dberkholz: yes, please |
| 06:44.29 | kblin | fortunately the results of the second part of my thesis are pretty good |
| 06:45.02 | kblin | especially considering I only started on that while waiting for lab results on my primary topic and was a bit bored :) |
| 06:45.12 | thebolt | kblin: haha, covered all bases ;) |
| 06:46.09 | Mkop | what's the second part? |
| 06:46.32 | dberkholz | Mkop: http://dev.gentoo.org/~dberkholz/personal/aconitase-overlay-of-rna-and-citrate-bound-forms.pse |
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| 06:47.05 | Mkop | hmm, some links are not meant to be clicked |
| 06:47.15 | Mkop | they are meant to be wgot |
| 06:48.00 | thebolt | Mkop: :) |
| 06:48.47 | kblin | Mkop: the RNA binding activity for these aconitases |
| 06:49.09 | dberkholz | Mkop: basically you'll see all kinds of green & blue helices that are the protein; the brown/blue one is rna; the yellow/orange blob is iron-sulfur cluster; the green/red blob is citrate; sidechains within 5 angstroms of the iron-sulfur cluster are also shown. |
| 06:50.27 | kblin | Mkop: prior to starting on this, my colleagues in microbiology could show that the streptomyces aconitases bind to the known eukaryote RNA sequence eukaryotic aconitase binds to |
| 06:50.58 | kblin | just that streptomycetes don't have the gene for e.g. ferritin |
| 06:51.18 | Mkop | dberkholz: how is the RNA positioned in there? randomly, or intentionally? |
| 06:51.19 | dberkholz | try frataxin instead |
| 06:51.31 | dberkholz | Mkop: it's position as it was bound in the experimentally determined structure |
| 06:51.43 | kblin | the overlay isn't too good, I guess |
| 06:51.56 | dberkholz | it's like a 3.5 A rmsd |
| 06:52.03 | kblin | in order to bind RNA, aconitase opens up around 60 A |
| 06:52.06 | kblin | oh, not too bad then |
| 06:52.15 | dberkholz | that's core regions |
| 06:52.21 | Mkop | how much is 60 Angstroms? |
| 06:52.21 | kblin | well, true |
| 06:52.57 | kblin | Mkop: in terms of structural alignments, a lot :) |
| 06:53.08 | Mkop | how wide is the whole protein? |
| 06:53.51 | Mkop | and, how do I get pymod to display the whole thing with a space-filling model? |
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| 06:53.56 | kblin | good question, didn't see that mentioned in a paper |
| 06:54.01 | dberkholz | you can see just looking at it that the overlay is reasonable but not great; the right side looks pretty ok, the left side is way off because of the domain shift |
| 06:54.04 | Mkop | not because it's any clearer that way, it's just cooler |
| 06:54.25 | dberkholz | Mkop: on the right, next to "all", click S then spheres |
| 06:54.27 | Mkop | dberkholz: what's your background in genetics/molecular biology? |
| 06:54.37 | dberkholz | or click surface, if you prefer |
| 06:54.50 | kblin | dberkholz: yeah, that's domains 1-3 staying the same, while domain 4 opens |
| 06:55.01 | Mkop | hehe |
| 06:55.06 | kblin | dberkholz: the same happens during the catalytic activity to allow substrate access |
| 06:55.09 | Mkop | you can't see anything with it on spheres |
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| 06:55.29 | dberkholz | yep, proteins are packed pretty tight. |
| 06:56.14 | kblin | so one of the things I actually found was that bacteria seem to be a bit more relaxed as to what the RNA pattern can look like |
| 06:56.21 | dberkholz | Mkop: structural biology |
| 06:56.33 | Mkop | is that what you do as your day job? |
| 06:56.42 | Mkop | and gentoo is your hobby? |
| 06:56.47 | dberkholz | yep |
| 06:57.02 | Mkop | what is structural biology? |
| 06:57.16 | Mkop | this kind of stuff? |
| 06:57.19 | dberkholz | basically using the structure to figure out the function, and explain known functions |
| 06:57.51 | Mkop | specifically of proteins? |
| 06:57.53 | dberkholz | to do so, you of course need to determine the structure somehow, and that's part of it too |
| 06:58.05 | dberkholz | macromolecules, which includes proteins, dna, rna, etc. |
| 06:58.18 | Mkop | DNA has functions? |
| 06:58.25 | Mkop | I know RNA does sometimes |
| 06:58.34 | dberkholz | depends on your definition of function |
| 06:58.43 | Mkop | I mean other than its primary role as data storage |
| 06:58.45 | Mkop | "data" |
| 06:58.56 | Mkop | there's also packing, etc. for DNA |
| 06:59.03 | dberkholz | does it catalyze chemical reactions? not that i know of |
| 06:59.17 | dberkholz | neither do lots of proteins though. |
| 06:59.25 | kblin | anyway, off to university for today, bbiaf |
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| 07:00.26 | Mkop | seeya, kblin |
| 07:00.30 | Mkop | this was very interesting |
| 07:00.31 | Mkop | thanks |
| 07:00.54 | Mkop | dberkholz: do you work in academia? |
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| 07:02.48 | thebolt | Mkop: as far as i can tell with that kind of area you either work in academia or pharmasutical company.. not much else to do ;) |
| 07:03.31 | Mkop | yeah, either in academics, or in industry research. my question is which |
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| 07:30.07 | kblin | re |
| 07:30.33 | thebolt | wb;) |
| 07:31.33 | kblin | dberkholz: I didn't find any iron-regulation-related genes with an IRE-like pattern in streptomyces genomes (IRE pattern being the iron-response relement pattern bound by eukaryontic aconitase) |
| 07:35.17 | smtms | kblin, can you repeat what you've just said? :-P |
| 07:43.10 | kblin | smtms: sure |
| 07:43.18 | kblin | smts: I didn't find any iron-regulation-related genes with an IRE-like pattern in streptomyces genomes (IRE pattern being the iron-response relement pattern bound by eukaryontic aconitase) |
| 07:48.16 | kblin | :) |
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| 07:49.05 | r0bby | kblin: what are you majoring in? |
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| 11:16.32 | kblin | I'm reading too much groklaw |
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| 11:17.58 | kblin | proteins usually aren't in litigation, they are in ligation |
| 11:20.09 | straw | :P |
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| 11:28.13 | MaNI | unless they are american proteins |
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| 14:52.11 | ajuonline | i got a card with someone else's name |
| 14:52.13 | ajuonline | :/ |
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| 14:55.37 | thiago_home | notify google |
| 14:56.09 | summatusmentis | also, stop taking over other people's houses |
| 14:56.09 | ajuonline | did already |
| 14:56.32 | ajuonline | summatusmentis: i dont even know that guy! although he is an Indian student |
| 14:56.58 | summatusmentis | uh huh. you looked up all the Indian students addresses, and took over their houses |
| 14:57.08 | summatusmentis | it's clearly all part of your world domination plan |
| 14:57.33 | ajuonline | shhh! |
| 14:57.34 | ajuonline | <PROTECTED> |
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| 19:18.05 | alidnoC | Hi everybody! Is there any project for Education (primary schooling)? |
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| 19:22.20 | alidnoC | I have a project called Our Education based on the topic from United Nations called Universal Education. |
| 19:22.40 | Catfish_Man | alidnoC: SoC is already going on; the time for projects to apply was months ago |
| 19:22.44 | alidnoC | And I believe that it will really help millions around the world. |
| 19:22.53 | alidnoC | Oh, I see. |
| 19:23.01 | alidnoC | Is it every year, though? |
| 19:23.09 | Catfish_Man | no guarantees, but very likely |
| 19:23.20 | andguent | why should the deadline stop you from "helping millions around the world" |
| 19:23.24 | alidnoC | Okay, I understand. |
| 19:23.37 | alidnoC | Well, I'm not very good at programming. |
| 19:23.43 | alidnoC | So I would need help. |
| 19:24.05 | alidnoC | I know some of the C programming language, but I haven't really gotten to the advanced level, yet. |
| 19:24.13 | alidnoC | Do you understand? |
| 19:24.45 | alidnoC | I developed it for the Imagine Cup - Software Design competition this year. |
| 19:25.02 | alidnoC | But since I don't know programming very well yet...I had to quit the competition. |
| 19:26.40 | ecin | alidnoC: Never a better time to develop your skills, then. :) |
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| 19:27.09 | alidnoC | That's true. |
| 19:27.24 | alidnoC | ecin, what's your e-mail address? |
| 19:27.35 | alidnoC | I can e-mail you its business plan. |
| 19:27.39 | alidnoC | Maybe you like it, too. |
| 19:27.39 | ecin | Ironically, I think Image Cup had education as their theme last year or the one before. |
| 19:27.56 | alidnoC | In 2007, and this year. |
| 19:28.07 | alidnoC | This year are more than one topics. |
| 19:28.14 | alidnoC | Including Universal Education, etc. |
| 19:28.34 | andguent | business plan? |
| 19:28.36 | andguent | omg. get out |
| 19:28.47 | dzhus | lol |
| 19:30.21 | alidnoC | Well, its lay out is like a business plan. |
| 19:31.48 | alidnoC | A business plan doesn't always mean the real deal like you were thinking. |
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| 22:19.26 | gurkeee | is someone here good @ css? :-) |
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| 22:35.36 | allisterb_ | gurkee, sort of |
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| 23:00.31 | chx | http://twitter.com/chx1975/status/2082461518 wooohoo <3 Nokia tablets. |
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| 23:14.01 | ajuonline | chx: great! now send that to me ;) |
| 23:14.38 | chx | send what? |
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| 23:17.34 | ajuonline | chx: tablets :P |
| 23:18.17 | chx | um |
| 23:18.20 | chx | just bought it |
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