Expression, purification, characteristics and homology modeling of the HMGS from Streptococcus pneumoniae.
OBJECTIVE
To understand the molecular basis for a potential reaction mechanism and develop novel antibiotics with homology modeling for 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) synthase (HMGS).


METHODS
The genetic engineering technology and the composer module of SYBYL7.0 program were used, while the HMGS three-dimensional structure was analyzed by homology modeling.


RESULTS
The mvaS gene was cloned from Streptococcus pneumoniae and overexpressed in Escherichia coli from a pET28 vector. The expressed enzyme (about 46 kDa) was purified by affinity chromatography with a specific activity of 3.24 micromol/min/mg. Optimal conditions were pH 9.75 and 10 mmol/L MgCl2 at 37 degrees C. The V(max) and K(m) were 4.69 micromol/min/mg and 213 micromol/L respectively. The 3D model of S. pneumoniae HMGS was established based on structure template of HMGS of Enterococcus faecalis.


CONCLUSIONS
The structure of HMGS will facilitate the structure-based design of alternative drugs to cholesterol-lowering therapies or to novel antibiotics to the Gram-positive cocci, whereas the recombinant HMGS will prove useful for drug development against a different enzyme in the mevalonate pathway.
Ben
Ya-Li
YL
College of Life Science, Central China Normal University, Wuhan 430079, Hubei, China. b_y_l@jhun.edu.cn
Cui
Gu-Zhen
GZ
Li
Chen
C
Han
Rui
R
Zhang
Jie
J
Zhang
Qing-Ye
QY
Wan
Jian
J
Liu
De-Li
DL
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D006904:Hydroxymethylglutaryl-CoA Synthase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011487:Protein Conformation; D013296:Streptococcus pneumoniae / Q000201:enzymology* / Q000235:genetics
D006904:Hydroxymethylglutaryl-CoA Synthase