This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Transhydrogenase (TH) is a dimeric integral membrane enzyme in prokaryotes and eukaryotic mitochondria that couples proton translocation across a membrane with direct hydride transfer between NAD(H) and NADP(H), which are bound in distinct soluble domains. Crystal structures of the NAD(H) binding domain of TH from Rhodospirullum rubrum (domain I) and the NADP(H) binding domain of bovine mitochondrial TH (domain III) have been solved previously in this laboratory, at resolutions of 1.2 and 1.8 respectively. Crystals of the NADP(H) binding domain of R. rubrum (domain III) have been obtained, with NADP+ and NADPH bound, and native data sets have been collected previously at SSRL, at resolutions of 2.1 and 2.4 respectively. In both cases, the protein crystallizes in a hexagonal bipyramidal form (space group P6(1)(2)(2), with cell dimensions, a = b = 117 and c = 211 The objective of the proposed MAD experiment is to obtain direct experimental phases using a selenomethionyl derivative of R. rubrum TH domain III.
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