Surface-entropy reduction used in the crystallization of human choline acetyltransferase.

Ae-Ri Kim, Tomas Dobransky, R Jane Rylett, Brian H Shilton

Department of Biochemistry, University of Western Ontario, London, Ontario N6A 5C1, Canada.

Acta crystallographica. Section D, Biological crystallography 2005 Sep

filter terms:

Human choline acetyltransferase (ChAT) synthesizes the neurotransmitter acetylcholine (ACh) from choline and acetyl-CoA. A crystal structure of human ChAT has been a long-standing goal in the neuronal signalling field. Milligram quantities of pure ChAT can be purified [Kim et al. (2005), Protein Expr. Purif. 40, 107-117], but exhaustive crystallization efforts failed to produce any crystals suitable for high-resolution structural studies. To obtain high-quality crystals of human ChAT, a truncation was made in a large poorly conserved loop region and high-entropy side chains were removed from the surface of the protein. The resulting 'entropy-reduced' ChAT (MR = 68.1 kDa) crystallizes readily and reproducibly and the crystals diffract X-rays to approximately 2.2 A. The availability of these crystals will allow us to study the structure of human ChAT on its own as well as in complex with its substrates and inhibitor molecules, leading to a greater understanding of its catalytic mechanism and regulation.

Citation

Ae-Ri Kim, Tomas Dobransky, R Jane Rylett, Brian H Shilton. Surface-entropy reduction used in the crystallization of human choline acetyltransferase. Acta crystallographica. Section D, Biological crystallography. 2005 Sep;61(Pt 9):1306-10