[en] On the basis of steady-state kinetics, inhibition of the exocellular dd-carboxypeptidase-trans-peptidase of Streptomyces R61 by β-lactam antibiotics was competitive with regard to the donor substrate. However, the complexes formed between the Streptomyces R61 enzyme and various β-lactam antibiotics were relatively stable, exhibiting half-lives of 40 to 80 min at 37°C and neutral pH. During breakdown of the complexes the protein underwent reactivation, whereas the released antibiotic molecule was chemically altered. With [14C]benzylpenicillin, the released compound was neither benzylpenicillin nor benzylpenicilloic acid. The properties of the Streptomyces R61 enzyme β-lactam antibiotic complexes were compared with those of the complexes formed between the same antibiotics and either the membrane-bound transpeptidase from Streptomyces R61 or the exocellular dd-carboxypeptidase-transpeptidase of Streptomyces R39.