Purification of penicillin-binding protein 2 of Escherichia coli

Abstract

Penicillin-binding protein 2 (PBP-2) of E. coli K-12 was purified by covalent affinity chromatography using 6-aminopenicillanic acid covalently coupled to carboxymethyl-SEpharose (6-APA-CM Sepharose). Purification of PBP-2 was accomplished by prebinding the methoxy cephalosporin, cefoxitin, to the Triton X-100-solubilized PBPs of E. coli and then incubating the PBPs ith 6-APA-CM-Sepharose. Cefoxitin readily binds to all the E. coli PBPs except PBP-2 and, thus, in the presence of cefoxitin, only PBP-2 could bind to the 6-APA-CM-Sepharose. The purification of a mixture of al of the PBPs of E. coli by affinity chromatography is also described.