Cloning, expression, purification, crystallization and X-ray crystallographic analysis of (S)-3-hydroxybutyryl-CoA dehydrogenase from Clostridium butyricum.

ABSTRACT: (S)-3-Hydroxybutyryl-CoA dehydrogenase from Clostridium butyricum (CbHBD) is an enzyme that catalyzes the second step in the biosynthesis of n-butanol from acetyl-CoA by the reduction of acetoacetyl-CoA to 3-hydroxybutyryl-CoA. The CbHBD protein was crystallized using the hanging-drop vapour-diffusion method in the presence of 2?M ammonium sulfate, 0.1?M CAPS pH 10.5, 0.2?M lithium sulfate at 295?K. X-ray diffraction data were collected to a maximum resolution of 2.3?Å on a synchrotron beamline. The crystal belonged to space group R3, with unit-cell parameters a = b = 148.5, c = 201.6?Å. With four molecules per asymmetric unit, the crystal volume per unit protein weight (VM) is 3.52?Å(3)?Da(-1), which corresponds to a solvent content of approximately 65.04%. The structure was solved by the molecular-replacement method and refinement of the structure is in progress.

SUBMITTER: Kim EJ 

PROVIDER: S-EPMC3976069 | biostudies-literature | 2014 Apr

REPOSITORIES: biostudies-literature