Aug 302007
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.

Aug 282007
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.

Aug 272007
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.

Biological data filters

 biology  Comments Off on Biological data filters
Aug 222007
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.

To learn NMR…

 books, nmr  Comments Off on To learn NMR…
Aug 212007
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).