Archive for 'Point Mutationalists'

This is new (doi: 10.1038/4551149a). Nature, a culturally elite British media journal much like The Economist, has endorsed Barack Obama for President of the United States.

From that liberal rag, Nature:

Some will find strengths in McCain that they value more highly than the commitment to reasoned assessment that appeals in Obama. But all the signs are that the former seeks a narrower range of advice. Equally worrying is that he fails to educate himself on crucial matters; the attitude he has taken to economic policy over many years is at issue here. Either as a result of poor advice, or of advice inadequately considered, he frequently makes decisions that seem capricious or erratic. The most notable of these is his ill-considered choice of Sarah Palin, the Republican governor of Alaska, as running mate. Palin lacks the experience, and any outward sign of the capacity, to face the rigours of the presidency.
[...]
This journal does not have a vote, and does not claim any particular standing from which to instruct those who do. But if it did, it would cast its vote for Barack Obama

Some will recall this particularly alarming YeTube video of Sarah Palin failin’ horribly at science:

I find it frightening – nay, terrifying – that someone with this utter lack of interest and capability should be given a stage upon which to run for the second highest office in the nation. To have someone in office that doesn’t understand the importance of Drosophila research – nor to have the wherewithal to investigate what a collaborative research effort is – should not be allowed near or in the White House. This isn’t intellectual laziness – this is stupidity. Let’s make sure we are all quite clear on that.

That is why, quite some time ago, The Chem Blog officially endorsed Barack Obama. While it was intended as somewhat of a joke, I was quite serious in my intentions. Obama is the candidate we need to change how we have conducted ourselves in Iraq, to change how we tax our people, to change how we fund science, to change how our public officials conduct foreign diplomacy, to change how the world views us… everything this administration has done needs to be undone post-haste. Change is more than just a meme – and when Obama wins, it will be a mandate.

Well deserved, Roger Tsein, Osamu Shimomura and Martin Chalfie will share the prize for their work on GFP.  The work is seminal in many regards, not the least of which is because it makes shit glow and glowing is way cooler then Pd couplings.  It’s important to note that I think Tsein should STILL RESPOND TO EMAILS no matter how many stupid fucking prizes he wins.  But that’s fine.

GFP is a very interesting molecule with amazingly bizarre chemistry at its core.  Central to the protein is a fluorphore – a conjugated heteroaromatic cluster – that emits green light.  Roger’s first publication on the subject was in 1995, iirc, so it took him a little more than 10 years to snag the highest honor in sciences for his efforts.  The detailed mechanisms of the actual biosynthesis have been remarkably well done and so, it is not unusual that such a discovery, which took less than 10 years to go from the lab to glowing mammals, should take a little more than that to go from obscurity to a Nobel Prize.

Above is an interactive X-Ray structure of GFP (that image was captured from my computer. You CAN find the chromophore, but you have to look for it.) In any regard, I find a good crystal structure to be refreshing and enjoyable. So click on that image up there and load the structure! You’ll notice a ton of fucking water. I wish I were more of an expert at JMOL so I could just have the protein up there, but it’s a sufficiently intuitive program that you should be able to explore the structure without great confusion.

Let me start off by saying that I’m a fan of Sheng-Hsien Chiu, but not of his new approach to rotaxane synthesis: kitsch (ACIEE DOI: 10.1002/anie.200803056).

I’ll be the first to admit that sometimes supramolecular chemists come up with some strange and, well, superficially useless creations but, much like the synthesis of Brevetoxins, the reason to make some of these molecules isn’t so much that the target itself is useful but that they are fundamentally interesting in their complexity and creating them “expands the toolbox” of the chemist.  Point is, it’s academic shit, you know. Cerebral. Thinkin’ man’s stuff like Molecular Borromean rings (of which S.-H. Chiu is a coauthor on that Science paper). What’s the purpose? To be cool and for you to wonder how they assemble and ask a bunch of questions.  It’s mental masturbation at its finest and you can do it with CPK models.  How awesome is that?

What is Sheng-Hsien’s recent contribution? Unprecidented smallness.

Using 21C7 Crown ether and a dipropargylammonium salt, they created that threaded thing up there. This, you see, is a “pseudorotaxane” and their formation has been reported with benzo21C7.  To seal the deal they did a Diels-Alder reactions with 1,2,4,5-tetrazine to create pyridazine end groups. They say they did the reaction in a ball-mill (solvent free), but Fritz Vögtle tortured generations of graduate students to show that you can make rotaxanes through all kinds of fucked up methods, I know melting the solids together was one, surely grinding them was one of them too.  If not I appologise (to Sheng-Hsien Chiu).  Even if Vögtle hasn’t, Sheng-Hsien Chiu published the proceedure in Angew. in 2008.  Which means there’s really no new chemistry.

What’s odd is, had I read this in Chem Commun or Org Let, I may have found this mildly cute and done a post on it as a cute little reaction, but I find the fact that it is in Angewandte a painful stretch. Then the authors FORCED me to do a post on it by concluding with:

Like athletes striving to achieve Olympic ideals – citius (swifter), altius (higher), fortius (stronger) – chemists have a new challenge when it comes to synthesizing [2]rotaxanes: minimus (smallest)!

Seriously. Maybe I’m being a huge douchebag here – I probably am – but the cornball cheese factor is dialed to fucking 11.

Chi-Chieh Hsu, Nai-Chia Chen, Chien-Chen Lai, Yi-Hung Liu, Shie-Ming Peng, Sheng-Hsien Chiu (2008). Solvent-Free Synthesis of the Smallest Rotaxane Prepared to Date Angewandte Chemie International Edition, 47 (39), 7475-7478 DOI: 10.1002/anie.200803056

This isn’t C&E news concentrates, but I want to cram a few in that I missed over the last few months. I’ve read and loved and you’ll love these advances in the field of chemistry:

First: DOI: 10.1126/science.1152692. Fucking brilliant piece from the Baker lab representing state of the art fuckability of proteins. The title: “De Novo Computational Design of Retro-Aldol Enzymes” gives you a half assed idea of what’s actually going on. Where the typical “state of the art” to select for proteins that do specific chemistry is to essentially make a fuck-ton of proteins and run them on a specially designed column or do some Systematic evolution of ligands by exponential enrichment. The idea is NOT to go and design an active site of a protein and then cut out the active site of another protein and stick your active site in it to do the chemistry you want it to do. My brief description trivializes the difficulty inherent here and may overstate the utility, but it’s a monumental push forward for protein engeneering that relies not upon random insertions or specific point mutations but de novo design of an active site and then finding a protein that is compatible with it.

Second: DOI: 10.1088/1468-6996/9/1/014104. Sir Fraser Stoddart and Bill Goddard collaborated to create one of the most awesome ideas in supramolecular chemistry , which I’ve gushed over here. The paper details the attempt and successful creation of the “quasi-tristable [2]catenane.” Limitations exist in the system and it’s apparent that the design, as it stands, is unlikely to produce the desired results as an unfortunate overlap appears to exist between the absorption and CT bands of the various colored bits, but I’m still undaunted in my interest of a system like this.

Finally: DOI: 10.1002/anie.200800891. The prior post makes this next post a bit more apropos. Design of rotaxanes with strong binding interactions necessarily makes “shuttling” them around rather difficult but the synthesis of interlocked molecules nearly necessitates strong interaction between thread and macrocycle to assemble. A Catch 22, as it were. Leigh and Zerbetto have a rather clever hack which is slightly reminiscent of some of Vogtle’s (and Leigh’s own) work, where the macrocycle is used to direct the chemistry that is going to make the rotaxane in the first place, though they use it in a distinctly clever way by exploiting the copper mediated Cadiot–Chodkiewicz reaction. I have pretty high hopes for this sort of thinking in the design of molecular shuttles.

These are just a few of the articles I read and found very interesting. If I weren’t so goddamn busy I’d give literature more treatment and limit my fluff posts. But, it’s my blog, so blah.

Yes. That’s right. We may not need to clone cute little babies for their cells, but we’ll still need their foreskins. (You don’t need it, you can use any skin tissue, but the authors of the Science paper used dick skin, which was too good to pass up.) Anyway, you can’t toss a stick without hitting some jackass talking about the latest skin-cells to stem cells advancement, simultaneously published in the journals Cell (2007, doi:10.1016/j.cell.2007.11.019 link to free PDF) and Science (ScienceExpress article DOI: 10.1126/science.1151526) – if your institution doesn’t receive either Cell or access to ScienceExpress you should either shoot your librarian or find another institution.

So, we should all know know the story by now… Scientist infects mouse with Lentivirus vector which causes the insertion of four genes, reverting the cells to their former stemey state. This is neat, so the scientists repeat it with human tissue found on the surface of some human and they get stemey looking cells. Hooray! you say, they’ve cured cancer/ethical dilemmas forever now I can start smoking again. WRONG. Nothing has changed. They took skin, either chopped off fresh from baby pecker (the Science article) or from some poor sap’s face (Cell) and infected it with a virus.

To avoid shuffling between the papers, I’m going to state, here and now, that the Science paper contains valid data and they achieved the same conclusions and did a good job and it has it’s own useful flavor or whatever, but compared to the Cell paper, it’s second rate. It looks less thoughtfully conducted and like the paper was written on the school buss on the way to school. It’s also quite Spartan on the details. So, you can read the Science paper and giggle at the fact that they used human dick cells, (to which one fellow researcher quipped, foreskin is great!) but beyond that, the Cell paper contains more useful (and more in quantity) experiments and discussion. Thus I quote (from Cell):

In this study, we showed that iPS cells can be generated from adult HDF and other somatic cells by retroviral transduction of the same four transcription factors with mouse iPS cells, namely Oct3/4, Sox2, Klf4, and c-Myc. The established human iPS cells are similar to hES cells in many aspects, including morphology, proliferation, feederdependence, surface markers, gene expression, promoter activities, telomerase activities, in vitro differentiation, and teratoma formation. The four retroviruses are strongly silenced in human iPS cells, indicating that these cells are efficiently reprogrammed and do not depend on continuous expression of the transgenes for self renewal. [...] The four factors, however, could not induce human iPS cells when fibroblasts were kept under the culture condition for mouse ES cells after retroviral transduction. These data suggest that the fundamental transcriptional network governing pluripotency is common in human and mice, but extrinsic factors and signals maintaining pluripotency are unique for each species. [...] Deciphering of the mechanism by which the four factors induce pluripotency in somatic cells remains elusive.

Awesome and a mystery to boot. So far as anyone looking at them from the outside is concerned, these bad boys are the real thing and, best of all, they don’t look like they’re handicapped by requiring some continuously expressing crap to “silence” their former status as skin cell.

We found that each iPS clone contained three to six retroviral integrations for each factor. Thus, each clone had more than 20 retroviral integration sites in total, which may increase the risk of tumorigenesis. In the case of mouse iPS cells, [approx 20% of the] mice derived from iPS cells developed tumors, which were attributable, at least in part, to reactivation of the c-Myc retrovirus…

Shitty. This is where the canz0r that you may have read about in the NYT comes in. It’s not the fact that it’s a stem cell, it’s the fact that it’s still loaded with viral vector. This *could* be overcome using current techniques like a gene gun or, hopefully, in the future some newer method would eventually be worked work out. But 20% is horribly unacceptable (beyond unacceptable, really) even though it provides awesome opportunities for work NOW.

Human iPS cells, however, are not identical to hES cells: DNA microarray analyses detected differences between the two pluripotent stem cell lines. Further studies are essential to determine whether human iPS cells can replace hES in medical applications.

Cautionary. (and frankly unnerving). Those differences may be irrelevant, of course. But it would suck if that new heart that you’re growing there turned back into a ball of face skin when the doctors put it in. So all this talk of “replacing”embrionic stem cell research is a bunch of hyperbolic crap. It may or it may not, we don’t know yet. It’s fun to get one’s hopes up, but it sucks to be let down. As a responsible scientist, you should keep in mind that this may or may not work out for many reasons, that statement up there being one of them. So don’t go around thinking it’s safe to pull the plug on funding for cloning and embryonic stem cell research ’cause the champaign that’s been flowing in the media was uncorked way too early. There is also the concern that the general inefficiency of this method (at present) won’t be useful for people who need quick organ repair. This is also something that time will likely improve upon.

But I don’t want to leave you with statements that are too prosaic. I’ll leave you with one of the finer things of the paper – a quote from somewhere in the middle:

We next examined whether lineage-directed differentiation of human iPS cells could be induced by reported methods for hES cells. We seeded human iPS cells on PA6 feeder layer and maintained them under differentiation conditions for 2 weeks (Kawasaki et al., 2000). Cells spread drastically, and some neuronal structures were observed (Figure 6A). Immunocytochemistry detected cells positive for tyrosine hydroxylase and bIII tubulin in the culture (Figure 6B). PCR analysis revealed expression of dopaminergic neuron markers, such as aromatic-L-amino acid decarboxylase (AADC), member 3 (DAT), choline acetyltransferase (ChAT), and LIM homeobox transcription factor 1 beta (LMX1B), as well as another neuron marker, MAP2 [...] These data demonstrated that iPS cells could differentiate into neuronal cells, including dopaminergic neurons, by coculture with PA6 cells.

translation:

My assistant and I, drunken with madness, pulled a dish of living brain cells out of the incubator. “What shall we do?” I asked. I knew I had to do the unthinkable. We thus took our mutant stem cells, cultivated from the faces of other humans, and layered them amongst the nerve tissue. The newly adopted cells spread violently across the dish! Axions whipped too and fro smacking my assistant to the ground. I grabbed the Brain-Analyzer to measure its chemical signals and INDEED! The creature forming before me was nothing less than the human brain. As the point mutation machine began to light up and spit out all kinds of numbers and letters – AADC, DAT, ChAT, LIM, LMX1b the only thing my assistant and I could do was watch; watch in horror. The monster before us wasn’t going to let us leave. It wanted to kill us – and God save us for creating such an abomination. It was waiting for the right time… the monster was thinking… (probably about abortion and limiting our gun rights and homosexuals. ARRRGH why did we let our guard down when they said it didn’t involve killing helpless clone babies?!)

They turned skin cells into brain tissue. How fucking awesome is that? Read the Cell paper, it’ll be hard (what with all the goddamn abbreviations used in biology) but it’s a good read. (If you have time read the Science one too. It’s very short and also reasonably good with a thoughtful concluding statement on what you should set your expectations levels at.)