Crystallization and preliminary X-ray crystallographic analysis of 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase from Mycobacterium tuberculosis.
The enzymes of the shikimate pathway are attractive targets for new-generation antimicrobial agents. The first step of this pathway is catalysed by 3-deoxy-D-arabino-heptulosonate-7-phosphate (DAH7P) synthase and involves the condensation of phosphoenolpyruvate (PEP) and erythrose 4-phosphate (E4P) to form DAH7P. DAH7P synthases have been classified into two apparently evolutionarily unrelated types and whereas structural data have been obtained for the type I DAH7P synthases, no structural information is available for their type II counterparts. The type II DAH7P synthase from Mycobacterium tuberculosis was co-expressed as native and selenomethionine-substituted protein with the Escherichia coli chaperonins GroEL and GroES in E. coli, purified and crystallized. Native crystals of M. tuberculosis DAH7P synthase belong to space group P3(1)21 or P3(2)21 and diffract to 2.5 A, with unit-cell parameters a = b = 203.61, c = 66.39 A. There are either two or three molecules in the asymmetric unit. Multiwavelength anomalous diffraction (MAD) phasing using selenomethionine-substituted protein is currently under way.
Webby
Celia J
CJ
Centre for Structural Biology, Institute of Fundamental Sciences, Massey University, Private Bag 11-222, Palmerston North, New Zealand.
Lott
J Shaun
JS
Baker
Heather M
HM
Baker
Edward N
EN
Parker
Emily J
EJ
D010723:3-Deoxy-7-Phosphoheptulonate Synthase; D000446:Aldehyde-Lyases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D018835:Chaperonin 10 / Q000235:genetics; D018834:Chaperonin 60 / Q000235:genetics; D018833:Chaperonins / Q000235:genetics; D003001:Cloning, Molecular / Q000379:methods; D003460:Crystallization / Q000379:methods; D004926:Escherichia coli / Q000235:genetics; D009169:Mycobacterium tuberculosis / Q000201:enzymology*; D012645:Selenomethionine; D014961:X-Ray Diffraction
D018835:Chaperonin 10; D018834:Chaperonin 60; D012645:Selenomethionine; D010723:3-Deoxy-7-Phosphoheptulonate Synthase; D018833:Chaperonins; D000446:Aldehyde-Lyases