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Hello all, I have a structure of my protein at 1.3 Ang. There is a clear density in the binding site but I can't seem to figure out what the ligand is. The crystalization conditions contain PEG, sulfate, TMAO and acetate buffer. The protein was purified by a Ni-NTA column and went into phosphate buffer after that. The ligand appears is interacting with a histidine and a water atom and is most likely acetate. Pictures can be viewed at www.xtalmerski.tumblr.com. However, there seems to be an additional atom on the acetate, which would be the easy answer but the buffer was of the 99+% purity (admittedly from Sigma) so I doubt that enough proprionic acid would make it through to account for 100% ligand occupancy. I also looked at TMAO decomposition by NMR under the same conditions for twice as long as the time it took to grow crystals and there was no apparent decomposition (which is 95% by NMR of course). A possible precendent in the literature for TMAO rearrangement requires a homolytic decomposition to formaldehyde and then formation of the new product to get oxygen migration which may be a bit of a stretch to suggest as a new enzyme catalyzed reaction.... Plus, refinement with that product isn't perfect either as the nitrogen has too many electrons to be in the tertiary center to fit this density. So, has anyone seen or heard or can imagine a way to get an extra atom on acetate? Thanks for any help you can give. Matthew Merski Post-doc Shoichet Group UCSF