Lia Andrea, Dowle Adam, Taylor Chris, Santino Angelo, Roversi Pietro
Leicester Institute of Chemical and Structural Biology and Department of Molecular and Cell Biology, University of Leicester, Henry Wellcome Building, Lancaster Road, LE1 7HB, UK.
Institute of Sciences of Food Production, C.N.R. Unit of Lecce, ia Monteroni, Lecce, 73100, Italy.
Wellcome Open Res. 2020 Aug 25;5:114. doi: 10.12688/wellcomeopenres.15893.2. eCollection 2020.
: n-Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) catalyses the NAD -dependent oxidative phosphorylation of n-glyceraldehyde-3-phosphate to 1,3-diphospho-n-glycerate and its reverse reaction in glycolysis and gluconeogenesis. : Four distinct crystal structures of human n-Glyceraldehyde-3-phosphate dehydrogenase ( GAPDH) have been determined from protein purified from the supernatant of HEK293F human epithelial kidney cells. : X-ray crystallography and mass-spectrometry indicate that the catalytic cysteine of the protein ( GAPDH Cys152) is partially oxidised to cysteine S-sulfonic acid. The average occupancy for the Cys152-S-sulfonic acid modification over the 20 crystallographically independent copies of GAPDH across three of the crystal forms obtained is 0.31±0.17. : The modification induces no significant structural changes on the tetrameric enzyme, and only makes aspecific contacts to surface residues in the active site, in keeping with the hypothesis that the oxidising conditions of the secreted mammalian cell expression system result in GAPDH catalytic cysteine S-sulfonic acid modification and irreversible inactivation of the enzyme.
：甘油醛-3-磷酸脱氢酶（GAPDH）催化甘油醛-3-磷酸在NAD依赖下氧化磷酸化为1,3-二磷酸甘油酸，并在糖酵解和糖异生过程中催化其逆反应。：已从人胚肾细胞系HEK293F上清液中纯化的蛋白质测定了人甘油醛-3-磷酸脱氢酶（GAPDH）的四种不同晶体结构。：X射线晶体学和质谱分析表明，该蛋白质的催化半胱氨酸（GAPDH Cys152）部分氧化为半胱氨酸S-磺酸。在获得的三种晶体形式中，20个晶体学独立的GAPDH拷贝上Cys152-S-磺酸修饰的平均占有率为0.31±0.17。：这种修饰在四聚体酶上未引起明显的结构变化，仅与活性位点的表面残基进行非特异性接触，这与分泌型哺乳动物细胞表达系统的氧化条件导致GAPDH催化半胱氨酸S-磺酸修饰和酶的不可逆失活这一假设相符。
