Expression, purification, and biochemical characterization of Mycobacterium tuberculosis aspartate decarboxylase, PanD

被引：35

作者：

Chopra, S

Pai, H

Ranganathan, A

机构：

[1] Int Ctr Genet Engn & Biotechnol, Recombinant Gene Prod Grp, New Delhi 110067, India

[2] Natl Brain Res Ctr, Gurgaon, Haryana, India

来源：

PROTEIN EXPRESSION AND PURIFICATION | 2002年 / 25卷 / 03期

关键词：

D O I：

10.1016/S1046-5928(02)00039-6

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

Like all bacteria, Mycobacterium tuberculosis (Mtb) possesses the genes necessary for coenzyme A biosynthesis and metabolism. In the present work, the Mtb panD gene was PCR amplified, overexpressed, and purified by metal affinity chromatography. The recombinant Mtb panD was found to exist as a tetramer in solution. incubation of Mtb panD at 37degreesC for several hours resulted in a complete cleavage of the inactive (pi) form into the two subunits (alpha and beta). The cleavage was confirmed by Western blot analysis as well as by N-terminal sequencing. Cleaved Mtb panD was assayed for decarboxylase activity with L-aspartate as substrate. The kinetic parameters K-m and k(cat) were found to be 219 muM and 0.65s(-1), respectively. These results provide the means for further studies based on the identification of the Mtb panD as well as other components of pantothenate metabolism as potential drug targets. (C) 2002 Elsevier Science (USA). All rights reserved.

引用

页码：533 / 540

页数：8

共 17 条

[1] Crystal structure of aspartate decarboxylase at 2.2 Å resolution provides evidence for an ester in protein self-processing [J].