Understanding the acylation mechanisms of active‐site serine penicillin‐recognizing proteins: A molecular dynamics simulation study

Abstract

Beta-lactam antibiotics inhibit enzymes involved in the last step of peptidoglycan synthesis. These enzymes, also identified as penicillin-binding proteins (PBPs), form a long-lived acyl-enzyme complex with beta-lactams. Antibiotic resistance is mainly due to the production of beta-lactamases, which are enzymes that hydrolyze the antibiotics and so prevent them reaching and inactivating their targets, and to mutations of the PBPs that decrease their affinity for the antibiotics. In this study, we present a theoretical study of several penicillin-recognizing proteins complexed with various beta-lactam antibiotics. Hybrid quantum mechanical/molecular mechanical potentials in conjunction with molecular dynamics simulations have been performed to understand the role of several residues, and pK(a) calculations have also been done to determine their protonation state. We analyze the differences between the beta-lactamase TEM-1, the membrane-bound PBP2x of Streptococcus pneumoniae, and the soluble DD-transpeptidase of Streptomyces K15.

Crystal Structure of a Peptidoglycan Synthesis Regulatory Factor (PBP3) from Streptococcus pneumoniae

“…This is followed by protonation of the nitrogen of the ␤-lactam ring by the carboxylate and subsequently ring breakage (35). An alternative concerted mechanism is possible, whereby the hydrogen from Ser 119 is transferred to the nitrogen of the ␤-lactam ring at the same time as the hydrogen of Ser 56 is transferred to Ser 119 , and the former forms the acyl bond with the antibiotic (37). sPBP3* is exceptionally efficient in hydrolyzing the pseudo substrate S2d, over 20 times more active than the most efficient pneumococcal enzyme measured to date, PBP2x (Table II) The numbering scheme follows that of the longest sequence, the E. faecalis enzyme (and thus does not relate to the one employed in the description of the sPBP3* structure).…”

Substrate specificity of bacterial DD-peptidases (penicillin-binding proteins)

“…It should also be noted, however, that neither of these analogues contains a leaving group and thus presumably most likely resemble deacylation transition states, 32, as analyzed below. It is possible that the leaving group (D-alanine) carboxylate might also participate in catalysis of the acylation step, as has been suggested for b-lactamases [144,145] (although not generally accepted in that case) and considered for DD-peptidases [146]. (34).…”

“…Interestingly, a lower pH shoulder (pKa 6.1) is seen in the data of Zhang et al [150], which might represent the real active enzyme. Computational approaches to DD-peptidase mechanisms have also been reported [145,146,154,155].…”

Crystal Structure of the Bacillus subtilis Penicillin-binding Protein 4a, and its Complex with a Peptidoglycan Mimetic Peptide

“…Acylation has been widely discussed in the framework of serine β-lactamases and more briefly for PBPs. 13,25 The mechanism encompassing the role of the lysine 55 as the general base is the most likely. 4,[9][10][11] The lysine needs to be unprotonated to abstract the proton from the active serine and, as seen in many crystallographic structures of PBPs, the proximity of Lys55 to Ser52 supports this idea.…”