Expression, purification and crystallization of penicillin G acylase from Escherichia coli ATCC 11105.
The penicillin acylase gene (pac) from Escherichia coli ATCC 11105 was cloned into pUC 9 and the resulting vector (pUPA-9), when transformed into E. coli strain 5K, allowed the constitutive overproduction of mature penicillin acylase when grown at 28 degrees C. The enzyme was purified from the periplasmic fraction of E. coli pUPA-9 by hydrophobic interaction chromatography and anion exchange. Crystals of penicillin acylase were grown in batch using polyethylene glycol 8000 as a precipitant. The crystals (space group P1) diffracted to beyond 2.3 A.
Hunt
P D
PD
Department of Chemistry, University of York, UK.
Tolley
S P
SP
Ward
R J
RJ
Hill
C P
CP
Dodson
G G
GG
D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D009154:Mutation; D010399:Penicillin Amidase / Q000096:biosynthesis / Q000235:genetics*; D012995:Solubility; D014961:X-Ray Diffraction
D010399:Penicillin Amidase