CRYSTAL-STRUCTURE OF A BACTERIAL SIGNAL PEPTIDASE IN COMPLEX WITH A BETA-LACTAM INHIBITOR

The signal peptidase (SPase) from Escherichia coli is a membrane-bound endopeptidase with two amino-terminal transmembrane segments and a ca rboxy-terminal catalytic region which resides in the periplasmic space (1). SPase functions to release proteins that have been translocated i nto the inner membrane from the cell interior, by cleaving off their s ignal peptides(1). We report here the X-ray crystal structure of a cat alytically active soluble fragment of E. coli SPase (SPase Delta 2-75) (2,3). We have determined this structure at 1.9 Angstrom resolution in a complex with an inhibitor, a beta-lactam (5S,6S penem)(4,5), which is covalently bound as an acyl-enzyme intermediate to the gamma-oxygen of a serine residue at position 90, demonstrating that this residue a cts as the nucleophile in the hydrolytic mechanism of signal-peptide c leavage. The structure is consistent with the use by SPase of Lys 145 as a general base in the activation of the nucleophilic Ser90, explain s the specificity requirement at the signal-peptide cleavage site, and reveals a large exposed hydrophobic surface which could be a site for an intimate association with the membrane. As enzymes that are essent ial for cell viability, bacterial SPases present a feasible antibacter ial target(4-6): our determination of the SPase structure therefore pr ovides a template for the rational design of antibiotic compounds.