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January 22, 2010
Receptors, Moving and Shaking
I've written here before about how I used to think that I understood G-protein coupled receptors (GPCRs), but that time and experience have proven to me that I didn't know much of anything. One of the factors that's complicated that field is the realization that these receptors can interact with each other, forming dimers (or perhaps even larger assemblies) which presumably are there for some good reason, and can act differently from the classic monomeric form.
A neat paper has appeared in PNAS that gives us some quantitative numbers on this phenomenon, and some great pictures as well. What you're looking at is a good ol' CHO cell, transfected with muscarinic M1 receptors. Twenty years ago (gulp) I was cranking out compounds to tickle cell membranes of this exact type, among others. The receptors are visualized by a fluorescent ligand (telenzepine), and the existence of dimers can be inferred by the "double-intensity" spots shown in the inset.
With this kind of resolution and time scale, the UK team that did this work could watch the receptors wandering over the cell surface in real time. It's a classic random walk, as far as they can tell. Watching the cohort of high-intensity spots, they can see changes as they switch to lower-intensity monomers and back again. Over a two-second period, it appeared that about 81% of the tracks were monomers, 9% were dimers, and 3% changed over during the tracking. (The remaining 7% were impossible to assign with confidence, which makes me wonder what's lurking down there).
They refined the technique by using two differently-fluorescent forms of labeled telenzepine, labeling the cells in a 50/50 ratio, and watching what happens to the red, green, (and combined yellow) spots over time. It looks as if the receptor population is a steady-state mix of monomers and dimers, exchanging on a time scale of seconds. Of course, the question comes up of how different ligands might affect this process, and you could begin to answer that with different fluorescent species. But since the technique depends on having a low-off-rate species bound to the receptor in order to see it, some of the most interesting dynamic questions will have to wait. It's still very nice to actually see these things, though; it gives a medicinal chemist something to picture. . .
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