Two-step procedure for purification and separation of the essential penicillin-binding proteins PBP 1A and 1Bs of Escherichia coli

The penicillin-binding proteins PBP 1A and 1Bs are the essential murein pol ymerases of Escherichia coli. Purification of these membrane-bound bifuncti onal transglycosylase-transpeptidases was a major obstacle in studying the details of both enzymatic reactions. Hen we describe a simple, highly speci fic affinity chromatography method that takes advantage of the availability of the specific inhibitor of the transglycosylase site moenomycin A in ord er to enrich PBP 1A and 1Bs in one step from crude membrane preparations. S eparation of PBP 1A from PBP 1Bs is achieved in a second step employing cat ion exchange chromatography yielding enzymatically active native murein pol ymerases. (C) 2000 Federation of European Microbiological Societies. Publis hed by Elsevier Science B.V. All rights reserved.