Crystallization and preliminary X-ray diffraction analysis of domain-chimeric L-(2S,3S)-butanediol dehydrogenase.
A domain-chimeric L-2,3-butanediol dehydrogenase (chimera L-BDH), which was designed to possess both the S-configuration specificity of L-BDH and the stability of meso-BDH, was constructed by exchanging the respective domains of these two BDHs. However, chimera L-BDH possessed a lower enzymatic function than expected based on the two original enzymes. To elucidate the causes of the decreased stability and substrate specificity, crystallization of the protein was performed. Chimera L-BDH was purified to homogeneity via ammonium sulfate fractionation and three column-chromatography steps, and was crystallized using the hanging-drop vapour-diffusion method. The crystals belonged to space group C2221, diffracted synchrotron radiation to 1.58 Å resolution and were most likely to contain two molecules in the asymmetric unit.
Graduate School of Medical and Engineering Science, Department of Education, University of Yamanashi, Japan.
Tomohito
T
Shimegi
Graduate School of Medical and Engineering Science, Department of Education, University of Yamanashi, Japan.
Takuji
T
Ooyama
Graduate School of Medical and Engineering Science, Department of Education, University of Yamanashi, Japan.
Takashi
T
Ohtsuki
Division of Structural Biology, Institute for Protein Research, Osaka University, Japan.
Genji
G
Kurisu
Graduate School of Medical and Engineering Science, Department of Education, University of Yamanashi, Japan.
Masami
M
Kusunoki
Graduate School of Medical and Engineering Science, Department of Education, University of Yamanashi, Japan.
Sadaharu
S
Ui
D000429:Alcohol Oxidoreductases / Q000737:chemistry* / Q000378:metabolism; D003460:Crystallization / Q000379:methods*; D004926:Escherichia coli / Q000201:enzymology; D008958:Models, Molecular; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D013237:Stereoisomerism; D014961:X-Ray Diffraction / Q000379:methods*
D000429:Alcohol Oxidoreductases; C019801:butanediol dehydrogenase
domain chimera; l-2,3-butanediol dehydrogenase