Structural And Functional Study Of The Type Ⅱ NADH Dehydrogenase

NDH2 is a single component type II N ADH dehydrogenase and acting as the alternative protein of multisubunit complex I. It plays the same role as complex I in catalyzing electron transfer from NADH to ubiquinone in the respiration chain. Located in the matrix side of the mitochondrial, the yeast NDH2(Ndi1) has a vital role in keeping the homeostasis of NADH/N AD+ in the mitochondrial. Besides, it can be used in the gene therapy of human diseases with complex I defects. Mainly existed in the lower organisms especially the pathogenic microorganisms and no homologous proteins in human being contributed the NDH2 protein an attractive drug target.Here we report the 2.39 angstrom crystal structure of Ndi1 by X-ray crystallography which is also the first structure of NDH2, and Ndi1-Q(2.48 angstrom), Ndi1-NADH(2.26 angstrom), Ndi1-NADH-Q(2.52 angstrom) complex structures. Ndi1 is a homodimer in all the structures and sequence aliment proved that Ndi1 has a conserved C terminal domain which is important for the dimerization, membrane anchoring and substrate recognization. The following biochemistry and cell biology experiments also proved the importance of C terminal domain. Together with the EPR experiments we proposed a model of probably electron transfer pathway which will help us to better understand the electron transfer mechanism of Ndi1 protein.We also expressed 7 other NDH2 s from different organisms, most of which are pathogenic, to conduct structural study, drug screen and optimization. Finally we got some attractive inhibitors that could inhibit the NDH2 protein well which will facilitate the drug design towards the diseases caused by these microorganisms.In this study, we present a comprehensive and systematic research of the most representative Ndi1 protein with multiple disciplines in its structure, function and the electron transfer mechanism. The research in the NDH2 s of the pathogenic organisms strengthened our knowledge of the structure information. We finally got some effective inhibitors that shed light on the drug design of diseases caused by the pathogenic microorganisms.