Characterization of the mimivirus cyclophilin

An interesting article showed up today on the Articles in Press page at the Journal of Molecular Biology. Vu Thai, the most talented young crystallographer I have never wanted to punch in the face, and Elan Eisenmesser, a man of sharp wits and sharper dress, along with several collaborators, have performed a really extensive characterization of the cyclophilin encoded in the mimivirus genome. This is an intriguing study because the role of cyclophilins in viral life cycles is still very poorly understood, even though they are known to aid the infectivity of HIV and SARS.

Typically, these viruses do not encode their own cyclophilins -- this would be a pretty costly thing for such a small and limited organism -- but rather hijack them from the local cellular machinery. What still hasn't been cleared up is exactly why that happens. The catalytic activity of cyclophilins is to convert proline residues from a cis conformation, in which the C alpha atoms of sequential residues lie on the same side of the C'-N bond, to a trans conformation in which they are on the opposite side. It's been demonstrated that human cyclophilin A (hCypA) catalyzes this conversion on the HIV capsid. However, it's not known whether this is the key role and reason for its incorporation. hCypA is also known to mediate some cell-binding events for HIV, and it's hypothesized that this is the only reason for inclusion, and the cis-trans isomerization is incidental.

The mimivirus cyclophilin (mimicyp) has the potential to answer some of these questions. One of the keys here is that it has been carried along in the viral genome. Because viruses are such efficient parasites operating under strong selective pressure, it stands to reason that anything the virus considers important enough to carry along with it is critical to its survival. Granted, mimivirus is a bit of an odd duck, one of the largest viruses known. Additionally, its primary targets are amoebae, though mimivirus has also been known to cause pneumonia by direct attack of human cells. And indeed, Thai et al. find that mimicyp most closely resembles an amoeboid cyclophilin.

The results start to get progressively weirder from there. Firstly, mimicyp has a vanishingly low affinity for cyclosporin, the molecule that gave cyclophilins their name. Moreover, it doesn't appear to have catalytic activity towards a standard proline isomerase substrate at all. The sequence of the protein is missing key residues that typically interact with isomerization substrates and contribute to catalysis, and its putative active site is uncharacteristic of active cyclophilins. Mimicyp crystallizes with an unusual trimeric arrangement, and Elan has demonstrated that it also forms multimers in solution that can be dissociated by adding arginine. Nonetheless, mimicyp localizes to the outer capsid of the mimivirus and is thought to be critical to infectivity, though Thai et al. were unable to demonstrate this for infection of Acanthamoebae polyphaga.

So what we have here is a cyclophilin that is catalytically dead, forms strange multimer arrays, and can't yet be shown to be important for infectivity, that nonetheless is always carried along by mature virions, and is so critical to the success of the virus that it is carried along in the genome and expressed despite significant selective pressure in favor of exclusion, mutation, or silenced expression. Granted some of these pressures are alleviated by the sheer size of the genome and virus, but others (particularly expression) are not. Thai et al. suggest a number of reasons why mimicyp might be important, including charge neutralization of the capsid, mediation of entry and viral disassembly, or capsid-masking similar to that performed by hCypA for HIV. And, of course, the possibility exists that mimicyp has some other, as-yet-unidentified catalytic activity that is essential for mimivirus infectivity, or is only critical for infecting certain amoebae or cells.

One might also surmise that the ability of mimivirus to infect vastly different hosts to some degree relies on its carrying along this particular cyclophilin, which is probably quite dissimilar to any protein abundantly found in human cells. This only brings us back to the question of why mimicyp is so important in the first place.

Ultimately this paper does not reveal the precise reason why cyclophilins are used by viruses, and indeed it will probably do more to spur debate than end it. Nonetheless, Thai et al.'s research can be taken as adding weight to the proposition that the catalytic activity of cyclophilins on virions is incidental, or at least that its importance is an idiosyncratic feature of particular viruses. Mimicyp may also be our first introduction to a new class of cyclophilins, catalytically inactive, cyclosporin-free (cyclophobins? cyclomehs?), and multimeric. What it is that these cyclophilins do and why they are important will be an intriguing inquiry to follow.

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A bittersweet day in the Kern Lab

Yesterday Peter, one of our MD guys, left the lab after two years. He and his wife are returning to Sweden now that she has completed her study here. All of us will miss his insanely delicate orange-peeling procedure and his absurdly neat notebooks, though I believe we bid adieu to his segment of mini-meeting without any regret. According to Doro, Peter really "stepped it up" in these last two months and should have two papers when we lazy experimentalists get off our butts and finish those projects. So I guess Peter will be hounding Annette and I for those valuable papers.

Dorothee was especially nice and thoughtful, telling Peter how much she appreciated all his hard work. She had come back to town from the cape just to see him off, and even arranged a little party. Before he left, she gave him a little gift, and I really felt the emotion of the occasion.

All of us wish Peter the best, and hope he finds a good job in Stockholm. I suppose Annette and I will have to "step it up" in order to put some good publications in the pipeline for him.

Disclaimer: One of the above paragraphs consists only of lies.

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Automation for your nanodevice assembly

An interesting article appeared in JACS ASAP today from C.S. Hartley and J.S. Moore at UIUC on a clever way to direct the assembly of an asymmetric macrocycle. The approach relies on a simple idea to address a fundamental question about nanostructures, namely, how to make sure they assemble themselves as we would like. Given that protein design still isn't quite far enough along to create any product we desire, we have to rely on more conventional chemistry. The basics of the approach should be obvious from the image on the right (shamelessly stolen from the paper). Note the reactive groups on the lower side of these compounds; even without knowing the relevant chemistry it should be clear what mechanism for directed assembly is implied. Given that the smallest geometrically stable assembly consists of three units, and that entropy favors the creation of small assemblies and enthalpy dislikes dangling functional groups, it should be obvious that dumping all these into a reactive pot should mostly produce units that have a composition of 1-1-1 or 1-2-3.

This is in fact what the authors observe when they perform the experiment, though they find (perhaps surprisingly) that in mixtures of all three components the 1-2-3 macrocycle predominates. Glancing at the product it's clear that the macrocycles themselves will assemble into stacked arrays given the right conditions. If covalent chemistry is used to control this assembly, one has a rather simple method that could be used to construct fairly complex structures. Also, the yields from these reactions were encouragingly rich in the desired molecules, suggesting that only a little further optimization is necessary to produce effective scale-up.

Nothing in the paper is particularly earth-shattering, but then again, the cleverest answers often seem blazingly simple in hindsight. Hartley and Brown have come up with just such an answer here, in a commendably clear and readable paper.

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Bioscience is murder

While reading Jane Stemwedel's musings on the missing gray zone in animal-research discussions, it occurred to me that when taken as a philosophical position, rather than an emotional appeal, the animal-rights people are really demanding that all bioscience be ended. I don't mean to make the old slippery-slope argument here; rather, I mean to say that when viewed in the light of cold reason, rather than sloppy anthropomorphism, the demand to eliminate animal suffering boils down to a demand to end bioscience research. Although we biochemists investigate life, our tool is death, and we use it constantly.

My objection to the animal-rights crowd hinges on their attribution of equivalent values to human and animal life, or human and animal pain and suffering. If you are willing to accept degrees, to say that a certain amount of animal suffering is tolerable in order to alleviate human suffering, then I am not arguing directly against you. I run into trouble when the crowd says that no outcome justifies animal research, because this argument implies that no research should be performed at all.

Consider the case of the laboratory mouse. On the one hand, there is an intuitive appeal to the animal-rights argument in their case. "How would you feel," they might well ask, "if you were locked up in a cage all day and subjected to experiments?" And of course, most of us would not like it at all, but that isn't relevant because our likes and dislikes are psychological constructs for which it is not clear mice have any equivalent. We're relatively closely related to mice, and they have behavioral characteristics similar to our own, which leads us to empathize with them, or so it seems. But this is a false empathy; we empathize with the mouse as if it is a human being with human traits and psychology, but this is manifestly not the case. The human mother, even a fecund one, mourns over her dead infant; the mouse mother eats hers. A human behaving as the mouse does would be called depraved, but we do not call the mouse depraved because we recognize that it does not think or feel as a human does. The similarity of responses to stressful stimuli owes more to a coincidence of behavioral evolution than matched psyches. Arguments against research on the basis of perceived empathy are therefore largely empty.

Given that anthropomorphic empathy is a dead end, the next line on animals is that they have intrinsic rights, especially that no animal individual should be used in research that does not help that individual. This implies something else, namely that an animal must consent to be researched upon. As it is impossible for the animal to give consent, we must imagine conditions under which it might give consent and stipulate those as a precondition to research. But on what basis to we attribute these rights? What virtue of a living creature endows it with the right to self-determination?

The most natural response to this question is that a creature gains the right for self-determination by having the ability to weigh the consequences of its actions and choose wisely. Indeed, this is the attribution by which we typically operate, and why we allow children and the mentally infirm only limited degrees of self-determination. However, this is clearly not the attribution the animal-rights activists are using -- a mouse does not even have the ability to conceive of the possible consequences of research, much less weigh them. For the animal-rights activist, the simple fact of being alive gives rise to the right to self-determination.

But why then should we stop with mice? Untold millions of drosophila have been bred and destroyed in the name of genetic research, not to mention everything that's been done with nematodes. Sure, it's more difficult to relate their behavior to ours, but these creatures live and die in captivity, and exhibit stress responses during certain experiments. What property of a mouse means that it has self-determination and the flies and worms don't?

And we're still being biased even if we let them in. After all, what's so special about multicellular organisms? Why should they be the only ones with a right to self-determination? If all it takes to require consent is being alive, then E. coli, of which I have personally raised and destroyed billions in the pursuit of NMR dynamics data, qualify too. In fact, their tale is really gruesome when you think about it. They're subjected to extreme temperatures and such rapid changes in them that they ingest large chunks of foreign DNA (themselves the product of enormous bacterial slaughter). I feed them a starvation diet loaded with strange chemicals, get them so high on IPTG they start to produce one protein almost exclusively, and then once they're done I murder them by repeatedly freezing and thawing them before I shatter their bodies with sound waves or crush them to death.

So there we are -- bacteria have rights, too, at least if you accept the reasoning of the most extreme (philosophically) animal rights supporters. I could go on to make a case for cultured cell lines as an independent life form, but really I've already gutted molecular biology, any cell biology involving DNA manipulation or foreign proteins (think how many bacteria died to bring you that Pfu turbo), and all of biochemistry and structural biology. If we grant that animals have the right to self-determination, none of the biosciences can possibly survive the scrutiny. That's what animal-rights activists are demanding, whether they know it or not.

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The disingenue

The ingenue in fiction is a sweet young girl who because of her naïveté is often taken advantage of by more worldly members of the cast. Sometimes she is even duped into committing some crime or other offense and made a scapegoat by dastardly villains. It's an enduring theme in literature, enough so that it still has traction in modern society, though in an altered form. I call the many who make use of it disingenues, a word that has been used occasionally by others without definition. By it I mean someone who pretends to be a naïve innocent led astray by people who pretended to be her friends, with the intention of deflecting blame for some misdeed. More broadly, it could be taken to mean someone who is disingenuous generally (like, say, Alberto Gonzales).

The narrow meaning could be applied to any number of misbehaving celebrities. They blame "bad influences" around them and pledge to improve the company they keep so as not to wander down the garden path again. It's a cheap dodge, and one that devoted fans always buy. Thinking seriously, when Britney Spears or Lindsay Lohan go out to party, who's in charge of that entourage? They are. They wield the power and authority in their circles, and they make their own decisions. Having the wrong people around them enhanced their opportunity to make bad choices, but those "bad influences" didn't choose for them.

But let's not limit the definition to women; this is the 21st century, and gender balance is at work here in epic proportions. The greatest recent disingenue is in fact a man, Michael Vick, who until this very day made a dedicated effort to lay blame for his own failures and shortcomings on the people surrounding him. From the beginning of this dog-fighting debacle, his position was that he had placed ill-deserved trust in his relatives and acquaintances. He had given them a house and they had betrayed him by using it for these evil purposes. Of course this fanciful tale of Mike Vick, ingenue, eventually unraveled, leading to today's guilty plea. Encouragingly, today he also said, "Not for one second will I sit right here and point the finger and try to blame anybody else for my actions or what I've done." Maybe he has turned the page, but consider how his statement of fact went to great lengths to indicate that the most gruesome act -- the violent killing earlier this year of several dogs that were not "game enough" -- was a "collective effort" involving Vick but not solely carried out by him. This is not materially different from the statements of fact accompanying his cohorts' pleas, but the difference in language is telling; he is still trying to deflect blame.

Literary or not, the disingenue is here to stay. This line of defense is too successful to fade anytime soon. We always want to pity a poor celebrity led astray by the bad influences around him or her. But let us not forget that these people chose to associate with those influences, and moreover, that when those influences offered a turn onto the wrong road, Mike Vick and others like him chose to take it.

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The brass ring is tarnished

News this week from the world of science: impact factor doesn't really match up with research quality. A study by Brown and Ramaswamy of the University of Iowa in Acta Crystallographica D analyzed crystallographic structures that had been deposited in the PDB and came up with a generalized metric to describe several features of a structure's quality. They normalized it to the average quality of the PDB, and then set about to see what extraneous factors were the best predictors of quality.

There were several interesting results here. The first was that the quality of crystal structures has not generally improved over time, despite advances in technique and equipment. The authors attribute this to the increasing use of crystallography as merely part of an experiment, so that the focus is less on the quality of the structure as a whole and more on making the structure good enough to answer a specific question. One can also imagine that the democratization of crystallography has contributed to this effect. Early structures solved with more primitive techniques only led to a solution if the researcher was qualified and careful. As the tools have become better, more widely available and black-boxed, more people can do crystallography, but these tools have encouraged less-qualified individuals to model structures, and perhaps also encouraged sloppiness among the better-trained researchers. There is also the vacuum-cleaner effect to consider. As the tools for modeling structures have become more robust, individual crystallographers are expected to produce more structures. As a result, the time spent on each individual structure has declined, perhaps more so than the improvements in technology have justified.

Another important result is that structural genomics consortia generally produce slightly better structures than the PDB average. This is certainly reassuring to the government entities that have dumped millions of dollars into these initiatives, but it also bears thinking about why it is so. Certainly one reason for the improved quality is formalized screening and the highly trained people that are doing the structural genomics work. Moreover, these efforts are really focused on getting a good structure, without any direct emphasis on experimental utility. It stands to reason that if your goal is to get a good structure, then it is more likely that those structures you personally consider a success and therefore deposit in the PDB will be good structures. This assumes a certain competence on the part of the researchers, but this is a property that structural genomics researchers manifestly possess. An additional consideration, however, is that these consortia are explicitly oriented towards solving the structures of well-behaved proteins. The high degree of automation used by the consortia is not generally compatible with poorly-behaved proteins or any need to squeeze a model out of troublesome data. A willingness to leave troubled proteins alone is probably part of the success of the consortia in this regard.

The most troubling result from the paper came when structures were analyzed based on the journal of primary reference. Ramaswamy and Brown discovered that more crystal structures initially reported in high-impact journals like Nature, Cell, and Science had below-average quality than structures reported in lower-impact journals like Biochemistry, Proteins, and Eur. J. Biochem. Because the metrics used by Brown and Ramaswamy are not intrinsically sensitive to novelty, this cannot be blamed on the simple "newness" of structures reported to these high-impact journals. The authors of this study attribute their problematic finding primarily to the fact that the structures reported in these journals tend to be (as above) part of a paper, not the whole thing. Moreover, the reviewers for these papers may more often be something other than expert crystallographers, and even when they are, the extreme space constraints may prevent reporting in the paper of the relevant structure factors and raw data.


That the reviewers are important might also be supported by the data. Consider the figure above (my own creation from Table 5 of the paper). This graphs the aggregate quality score for a journal against the number of structures therein - remember that a lower aggregate score is better. There's clearly no linear correlation between quality (lower numbers are better) and the number of structures reported. But it is interesting to see that with two exceptions the 'bad' journals (positive quality score) have a number of structures between 100 and 1000. Inside this range the quality varies significantly but outside of it the quality is almost always good. Maybe this means nothing, but it also may mean that journals who have a significant stock in trade in protein crystal structures also have experienced editors and reviewers who know how to properly vet them. It's interesting to note that the only journal containing more than a thousand structures that has a positive score is Proceedings of the National Academy of Sciences, a journal which has a strange and inconsistent review policy. Similarly, journals where crystal structures are a rare event may have editors who react to them with caution and seek out expert reviewers. By contrast, in the middle range, no trend is discernible except a roughly inverse relationship between impact factor and structure quality.

The more disturbing implication, and one that the authors do not deeply address, is that an external property of novel structures causes them to be published with lower quality: namely, their very novelty makes quality more of an afterthought. This is not just the idea of rushing papers to publication causing trouble. Rather, I mean to say that the editors and reviewers of these papers are flat-out willing to accept lower quality of data in exchange for novelty and impact. In, for example, the recent high-profile pentaretraction by Geoffrey Chang, the reported features of the structures alone should have raised serious questions about publishing them, even without considering that they were contradicted by biochemical data. That these errors somehow did not rise to the level of alerting Chang to find the elementary error in his own software indicates that the reviewers and editors were as sloppy as Chang himself. And why? Because these structures were novel and potentially revolutionary. Good copy outweighed bad modeling.

Crystal structures have a useful feature in that their quality can to some degree be assessed quantitatively, without needing to ask subjective questions (say, whether a structure is consistent with some mechanistic model). That is, this kind of large-scale analysis of research quality is possible. The quality of structures in these journals isn't disastrous, but it is cause for concern. And it raises serious questions about other research published in these journals, data for which the quality is less quantifiable and objective. I won't say that Science and Nature are not to be trusted, but in light of this large-scale trend and recent data-falsification woes in other areas, it would be naive in the extreme to approach reports in these journals without a healthy skepticism.

Brown E.N., Ramaswamy S. "Quality of protein crystal structures" Acta. Cryst. D. 63 (2007) p. 941-950

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Biological data filters

I had a conversation this week with Annette about the structural ensemble versus individual structures that I'm still trying to coalesce into a fully-formed idea. The kernel of it is this: there is a dichotomy between the way we know that proteins act and the way we talk about their action. Proteins give rise to phenomenological effects as ensembles, but we discuss their states as individuals.

Consider a signaling molecule, say a member of a MAP kinase cascade (picture at right). A given protein can exist in either an inactive (A) or active (B) form. When active, the kinase phosphorylates some downstream target, otherwise it just sits in your cytoplasm taking a nap. Typically in this kind of system the active form of the kinase is also the phosphorylated form (red B). It's typical to say something like, "The kinase is activated by phosphorylation." At the same time we know from some of Dorothee's work with Dave Wemmer that certain bacterial proteins that get phosphorylated already sample their active conformations even before they are modified (blue B).

Even for systems where this kind of sampling hasn't been directly demonstrated it's reasonable to assume it takes place. After all, the active and inactive structures have the same amino acids to work with. Unless the phosphate group itself is a lynchpin of the new structure (perhaps by bridging two structural elements), then the active structure must be one the unmodified kinase can adopt. Naturally, we expect this structure to be higher in energy than the inactive state (so blue B is higher on our energy diagram than blue A), and that phosphorylation decreases the energy of the active state so that it is subsequently preferred (so red B is lower than blue A).

The implication of this is that, unless the unmodified active structure is much higher in energy than the inactive structure, some proportion of our kinase is active even when not phosphorylated. Perhaps this is as low as 1-2%, a fraction that's difficult to detect directly. Still, because enzymes are so efficient, this quantity may be significant. Or, for a single protein, we could say that it adopts an active form without phosphorylation 1-2% of the time. But we tend to talk about phosphorylation and other post-translational modifications as if they were switches, with phrases like "protein X is turned on by phosphorylation". The reality, though, is that the switch is less a matter of turning a protein on than of turning it on more.

This points to a reality far less clean and orderly than typically depicted in block schematics. Inappropriately active (i.e. active without modification) members of the various protein ensembles must give rise to a considerable amount of noise in biological information processes. The system must therefore have some way to distinguish the signal from the noise that's more than just the binary on/off typically depicted and discussed. These filters could take several forms -- for instance, the kinase of our kinase may mediate the interaction between our kinase and its target, though in this case inappropriate activation of the MAPKK could still give rise to signaling noise. Alternately, the phosphate could mediate the kinase - target interaction. Or the cell could simply have an inefficient signaling system, so that multiple nearly-simultaneous signaling events are necessary to activate a response.

Is this point important? Maybe and maybe not. Most of our experiments can only access the behavior of ensembles, so the ensemble nature of protein action is not likely to lead us astray. But as single-molecule studies become more popular it may be important to keep the ensemble perspective in mind so as not to be confused by their results. Moreover, a conceptually accurate picture of cellular signaling and regulation will require us to keep this feature in mind.

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To learn NMR...

I picked up James Keeler's Understanding NMR Spectroscopy because of a teaching dilemma. Students who come to an NMR lab often want to "learn NMR", though of course this is not really possible in a 2-3 month rotation. They can at least get started in NMR, but in order to do this effectively they need a resource to study and discuss with whomever has charge of them in the lab. I've had trouble finding an appropriate book for this task. High-Resolution NMR Techniques in Organic Chemistry, originally by Derome and now reincarnated by Claridge, is a fine introduction for general NMR study but is not at all oriented towards biomolecules and relies heavily on the vector representation that doesn't always help a student understand techniques such as HMQC or HSQC. Protein NMR Spectroscopy, by Cavanagh, Fairbrother, Palmer, and Skelton is an excellent resource for the advanced student, and has just come out with the long-awaited new edition, but the pages of mathematics and occasionally obscure language are really too intimidating for beginners.

Understanding NMR Spectroscopy is, I think, the resolution of this dilemma. Keeler's text is clear, describing the physical basis of NMR in a straightforward way that should work for just about any student. He handles the necessary quantum mechanics and operator representations with a deft touch that makes their mathematical derivations clear without producing an intimidating morass of equations. Naturally, some detail and rigor is swept under the rug in this approach, and an advancing student will want the Cavanagh book or Levitt's Spin Dynamics to get a firmer grasp of the nuts and bolts, but as an introduction to the theoretical underpinnings Understanding NMR Spectroscopy is superb. Keeler's explanation of relaxation processes is also excellent, and includes perhaps the best physical description of T2 relaxation I have ever read. The book also includes a useful little chapter on the workings of an NMR spectrometer that, while nothing special on its own, is also a good resource for an early-career grad student or rotator. Exercises at the end of each chapter can also be a good teaching tool (although, since the answers are available at spectroscopyNOW, not appropriate for a course).

Although the book is not explicitly oriented towards biomolecular NMR, it has a strong focus on heteronuclear experiments that ensures the information presented is appropriate for students interested in biomolecules.

I can't praise this book without reservation, however. Some topics that might be considered important are glossed over or skipped entirely -- chemical exchange, for example is barely mentioned, and REX not at all. Residual dipolar couplings are not discussed, and the angular dependence of the dipolar interaction is only skimmed. Chapters 10 and 11 are poorly structured and include inadequate and possibly confusing discussions of raising and lowering operators, coherence order, and coherence transfer pathways and diagrams. The mentor will need to take an active hand in explaining just what is going on in these sections.

That said, I think that Understanding NMR Spectroscopy will be an excellent book for grad students just starting out in biomolecular NMR or possibly rotating students who want a glimpse of the nuts and bolts of NMR theory. The gap between Derome/Claridge and Cavanagh has been pretty neatly filled by this affordable little volume ($40 at Amazon).

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"Jealousy among men is a terrible thing..."

Last night I rented and watched Taboo, or Gohatto if you use the original Japanese title. I don't think I would have found it on my own, but the Netflix recommendation algorithm spat it out and I gave it a try. Taboo is a fascinating, well-acted gay samurai mystery art film, and you cannot be any more surprised by reading that than I was by writing it.

The conceit of the film is that an exceptionally pretty boy named Kano Sozaburo joins a samurai military unit towards the end of the Shogunate (the politics of this era play a minor, but substantive role in the story). Many of the men become infatuated with him, and their jealousy starts to tear the unit apart until a bloody episode that ends the film. Beat Takeshi stars as the unit's second-in-command Hijikata Toshizo, who is the one that has to deal with the problems Kano creates. These are many, as Kano seems to catch absolutely everyone's eye, including the commander, at least one lieutenant, and even perhaps Hijikata himself.

This is not the best movie I've ever seen by any standard. The plot seems to come and go at times, and the tension builds only unevenly towards the final confrontation. While the movie centers on Kano little is done to give the viewer a real handle on his character. He blows hot and cold, and Matsuda Ryuhei at times just doesn't seem equal to his task. Takeshi does more with less; Hijikata is much easier to grasp, even though he has his own set of unresolved questions. In the end the viewer is left to resolve most of the movie's central mysteries.

The directing is very interesting. Oshima Nagisa's camera tries to let action speak for itself, which works splendidly in some scenes (Kano's embarrassing kendo bout against his lover) and not so well in others (an extended entrance by a geisha). As is often the case, color features prominently in the film (Kano's frequent wearing of white against the black of the militia's livery), but it bears thinking about what the colors mean in the cultural context.

Some cultural context may also be needed for westerners watching the film. An American might find it strange that all these men are attracted to Kano, who seems by our standards pale and rather pinched. The seeming lack of romance in the love scenes (there is one sex scene, but barely anything is shown) may also seem a little odd. These aren't flaws but rather important components of the historical and cultural setting, ultimately helping to build a fairly accurate picture of this world.

Taboo isn't easy to understand and just isn't for some people. I don't mean this pejoratively; you aren't a better person if it is for you. It's the strangest case ever made against gays in the military -- beware, all the straight men might become infatuated! And yet, though some of the plot seems totally alien, it works, even the slightly strained ending. Beat Takeshi is to be credited for this, as he grounds a film that could have easily flown off into space. Sakagami Jiro, Takeda Shinji, and a few others in the supporting cast also do great work, propping up (in my opinion) Matsuda, who just doesn't seem to have quite enough poison in his eyes for this role. If you think a gay samurai mystery art film might be for you, then try to get your hands on it, at least for a rental.

I also watched Hot Fuzz; all you need to know about this film is that it is very funny and you should rent it.

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Leverage

Man, there is a lot of chatter about Mike Vick these days. I used to think he wasn't quite as big a punk as his brother, but apparently family ties run deep. The talking heads all seem to expect a 1-year sentence at minimum, but I'm not so sure. Yeah, the Feds could give Vick all that jail time and make an example of him, but to me that seems like squandering their advantage. The sports guys have it right when they say that Vick's career can't really take even a year off -- he's an athletic freak, not a football mind, and without the constant exposure to field conditions his already-deficient technique could take an irreparable hit. If he spends a year in jail, that means he'll miss at least two seasons, and if that happens he may have no choice other than to come back as a running back. So basically the Feds have him over a barrel. If they hit him with even a fairly light sentence, his career might well be over.

But all that gets the Feds is Bad Newz Kennels. Wouldn't it make more sense to slap his wrist in exchange for having him roll over on all those guys his dogs fought against? There was a lot of talk early on in the investigation about how Mike Vick was a major player in the dogfighting world. The Feds can use that knowledge -- after all, the world of dogfighting has been notoriously difficult to breach -- and I think they might dangle a big fat carrot in front of Vick to get it. I think that maybe his plea deal will involve a 6-month sentence at Club Fed, with another 3-month commitment in front of a grand jury. Now if only somebody would throw Pacman Jones in jail...

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The Persistence of Memory

There's a neat article in today's Science that could creep you out. Shema, Sacktor, and Dudai demonstrate that an inhibitor of protein kinase M zeta, called ZIP, apparently erases the long-term memories of rats. Rats presented with a novel taste that makes them sick subsequently avoid water with that taste (this is called Conditioned Taste Aversion). If the rats are treated with ZIP after the initial conditioning, they lose this aversion, even if treated with ZIP up to 25 days after the initial conditioning. ZIP treatment prior to conditioning has no effect. The implication is that ZIP has erased the memory that gives rise to aversion. Moreover, this effect depends on where in the brain the ZIP is infused (hippocampal infusion has no effect), suggesting that there is some possibility of controlling which memories are removed.

For people suffering from post-traumatic stress disorder or similar problems this could be the beginning of a promising therapeutic avenue. Of course, for those of a more imaginative bent this holds the promise of being a frightening brainwashing tool.

Check out the research article and commentary in the August 17 issue of Science.

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Small-molecule allosteric catalyst

An interesting pair of articles showed up this week, both from Chad Mirkin's lab at the Nanotechnology Center of Northwestern University. You can find them both in the August 22 edition of JACS. The essence of these articles is that Mirkin's group has created a group of macrocyclic catalysts that can be allosterically regulated by small ligands. The basic principle is shown below (figure taken from the second paper):

The catalytic molecule has a small intrinsic ability to catalyze the chemical conversion of the small molecules, as shown on the left. The addition of the "Analytes" in this case Cl- and CO, displaces the sulfur coordinating groups that are weakly bound to the rhodium or copper metal centers. Removing the sulfur coordination allows the molecule to undergo a conformational change, opening up to adopt a new structure that is vastly more effective as a catalyst. This mimics the allosteric regulation of protein catalysts and binding proteins by ligands.

The approach is a pretty clever bit of molecular engineering, and an important step forward for nanotechnology. Though it remains to be seen how well these molecules survive under field conditions (reducing/oxidizing environments, complex solutions), they have a feature that will be absolutely essential for functional nanomachines, namely that they can be regulated. The ability to turn our microscopic tools on and off will be an important part of producing useful nanomachines, to say nothing of avoiding the "gray goo" of sci-fi nightmares.

If you're interested, check out these articles:
Kuwabara, J. Stern, C.L., and Mirkin, C.A. "A Coordination Chemistry Approach to a Multieffector Enzyme Mimic", J. Am. Chem. Soc. 129 (2007) p. 10074-10075.

Masar III, M.S., Gianneschi, N.C., Oliveri, C.G., Stern, C.L., Nguyen, S.T., and Mirkin, C.A. "Allosterically Regulated Supramolecular Catalysis of Acyl Transfer Reactions for Signal Amplification and Detection of Small Molecules" J. Am. Chem. Soc. 129 (2007) p. 10149-10158.

I recommend the first article for lay readers, and the second for the hardcore.

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