Expression, purification, crystallization and preliminary X-ray crystallographic analysis of L-lactate dehydrogenase and its H171C mutant from Bacillus subtilis
Journal Article
·
· Acta Crystallographica Section F: Structural Biology and Crystallization Communications
- MSU
L-Lactate dehydrogenase (LDH) is an important enzyme involved in the last step of glycolysis that catalyzes the reversible conversion of pyruvate to L-lactate with the simultaneous oxidation of NADH to NAD{sup +}. In this study, wild-type LDH from Bacillus subtilis (BsLDH-WT) and the H171C mutant (BsLDH-H171C) were expressed in Escherichia coli and purified to near-homogeneity. BsLDH-WT was crystallized in the presence of fructose 1,6-bisphosphate (FBP) and NAD{sup +} and the crystal diffracted to 2.38 {angstrom} resolution. The crystal belonged to space group P3, with unit-cell parameters a = b = 171.04, c = 96.27 {angstrom}. BsLDH-H171C was also crystallized as the apoenzyme and in complex with NAD{sup +}, and data sets were collected to 2.20 and 2.49 {angstrom} resolution, respectively. Both BsLDH-H171C crystals belonged to space group P3, with unit-cell parameters a = b = 133.41, c = 99.34 {angstrom} and a = b = 133.43, c = 99.09 {angstrom}, respectively. Tetramers were observed in the asymmetric units of all three crystals.
- Research Organization:
- Advanced Photon Source (APS), Argonne National Laboratory (ANL), Argonne, IL (US)
- Sponsoring Organization:
- DOE - BASIC ENERGY SCIENCESOTHER U.S. STATES
- OSTI ID:
- 1049536
- Journal Information:
- Acta Crystallographica Section F: Structural Biology and Crystallization Communications, Journal Name: Acta Crystallographica Section F: Structural Biology and Crystallization Communications Journal Issue: 1 Vol. 68
- Country of Publication:
- United States
- Language:
- ENGLISH
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