A crystallographic comparison between mutated glyceraldehyde-3-phosphate dehydrogenases from Bacillus stearothermophilus complexed with either NAD+ or NADP+

Journal of Molecular Biology
C DidierjeanA Aubry

Abstract

Mutations have been introduced in the cytosolic glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from Bacillus stearothermophilus in order to convert its cofactor selectivity from a specificity towards NAD into a preference for NADP. In the B-S mutant, five mutations (L33T, T34G, D35G, L187A, P188S) were selected on the basis of a sequence alignment with NADP-dependent chloroplastic GAPDHs. In the D32G-S mutant, two of the five mutations mentioned above (L187A, P188S) have been used in combination with another one designed from electrostatic considerations (D32G). Both mutants exhibit a dual-cofactor selectivity at the advantage of either NAD (B-S) or NADP (D32G-S). In order to analyse the cofactor-binding site plasticity at the molecular level, crystal structures of these mutants have been solved, when complexed with either NAD+ (D32G-Sn, resolution 2.5 A, R = 13.9%; B-Sn, 2.45 A, 19.3%) or NADP+ (D32G-Sp, 2.2 A, 19.2%; B-Sp, 2.5 A, 14.4%). The four refined models are very similar to that of the wild-type GAPDH and as expected resemble more closely the holo form than the apo form. In the B-S mutant, the wild-type low affinity for NADP+ seems to be essentially retained because of repulsive electrostatic contacts between the ext...Continue Reading

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Citations

Aug 10, 2011·Proceedings of the National Academy of Sciences of the United States of America·Steven D BrownLee R Lynd
Jan 1, 2014·Journal of Bioinformatics and Computational Biology·Meiyappan LakshmananDong-Yup Lee
Feb 16, 2007·Journal of Molecular Biology·Yoshiaki YasutakeTomohiro Tamura
Jan 21, 2001·The Journal of Biological Chemistry·M MedinaC Gomez-Moreno
Oct 14, 2016·Chembiochem : a European Journal of Chemical Biology·Andy BeierUwe T Bornscheuer
Dec 14, 2001·The Journal of Biological Chemistry·Fabrice MoucheNicolas Boisset
Apr 5, 2014·Acta Crystallographica. Section D, Biological Crystallography·Kudigana J PampaRavishankar Vittal Rai
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Jul 11, 2006·Biochemical and Biophysical Research Communications·Takeo TomitaMakoto Nishiyama
Mar 31, 2004·Biochemistry·Limei Tao, Ann M English

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