Synthesis and Kinetic Analysis of Two Conformationally Restricted Peptide Substrates of Escherichia coli Penicillin-Binding Protein 5.

Escherichia coli PBP5 (penicillin-binding protein 5) is a dd-carboxypeptidase involved in bacterial cell wall maturation. Beyond the C-terminal d-alanyl-d-alanine moiety, PBP5, like the essential high-molecular mass PBPs, has little specificity for other elements of peptidoglycan structure, at least as elicited in vitro by small peptidoglycan fragments. On the basis of the crystal structure of a stem pentapeptide derivative noncovalently bound to E. coli PBP6 (Protein Data Bank entry 3ITB ), closely similar in structure to PBP5, we have modeled a pentapeptide structure at the active site of PBP5. Because the two termini of the pentapeptide are directed into solution in the PBP6 crystal structure, we then modeled a 19-membered cyclic peptide analogue by cross-linking the terminal amines by succinylation. An analogous smaller, 17-membered cyclic peptide, in which the l-lysine of the original was replaced by l-diaminobutyric acid, could also be modeled into the active site. We anticipated that, just as the reactivity of stem peptide fragments of peptidoglycan with PBPs in vivo may be entropically enhanced by immobilization in the polymer, so too would that of our cyclic peptides with respect to their acyclic analogues in vitro. This paper describes the synthesis of the peptides described above that were required to examine this hypothesis and presents an analysis of their structures and reaction kinetics with PBP5.