This study deals with the interaction of nocardicin A with Escherichia coli penicillin-binding proteins. Competition experiments with two different isotopically labelled beta-lactams indicated that nocardicin A interacts with penicillin-binding proteins 1a, 1b, 2 and 4 in intact cells. Binding of nocardicin A to the penicillin-binding proteins was abolished, or greatly reduced, when the assays were carried out with purified cell envelopes. Important differences between the binding patterns of benzyl[14C]penicillin to intact cells and to purified cell envelopes were also observed. These results suggest that the interaction of beta-lactam antibiotics with their target proteins depends to a very great extent on the state of the cell envelope as a whole.