N-Acetyl-L-glutamate kinase from Escherichia coli:: cloning of the gene, purification and crystallization of the recombinant enzyme and preliminary X-ray analysis of the free and ligand-bound forms

被引:14
作者
Gil, F
Ramón-Maiques, S
Marina, A
Fita, I
Rubio, V
机构
[1] CSIC, Inst Biomed Valencia, E-46010 Valencia, Spain
[2] CSIC, IBMB, Barcelona, Spain
来源
ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY | 1999年 / 55卷
关键词
D O I
10.1107/S0907444999005351
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
The gene for Escherichia coli N-acetyl-L-glutamate kinase (NAGK) was cloned in a plasmid and expressed in E. coli, allowing enzyme purification in three steps. NAGK exhibits high specific activity (1.1 mu mol s(-1) mg(-1)), lacks Met1 and forms dimers (shown by crosslinking). Crystals of unliganded NAGK diffract to 2 Angstrom and belong to space group P6(1)22 or its enantiomorph P6(5)22 (unit-cell parameters a = b = 78.6, c = 278.0 Angstrom) with two monomers in the asymmetric unit. Crystals of NAGK with acetylglutamate and the ATP analogue AMPPNP diffract to 1.8 Angstrom and belong to space group C222(1) (unit-cell parameters a = 60.0, b = 71.9, c = 107.4 Angstrom), with one monomer in the asymmetric unit. NAGK crystallization will allow the determination of proposed structural similarities to carbamate kinase.
引用
收藏
页码:1350 / 1352
页数:3
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