PMID:10967102
| Citation |
Davies, C, White, SW and Nicholas, RA (2001) Crystal structure of a deacylation-defective mutant of penicillin-binding protein 5 at 2.3-A resolution. J. Biol. Chem. 276:616-23 |
|---|---|
| Abstract |
Penicillin-binding protein 5 (PBP 5) of Escherichia coli functions as a d-alanine carboxypeptidase, cleaving the C-terminal d-alanine residue from cell wall peptides. Like all PBPs, PBP 5 forms a covalent acyl-enzyme complex with beta-lactam antibiotics; however, PBP 5 is distinguished by its high rate of deacylation of the acyl-enzyme complex (t(12) approximately 9 min). A Gly-105 --> Asp mutation in PBP 5 markedly impairs this beta-lactamase activity (deacylation), with only minor effects on acylation, and promotes accumulation of a covalent complex with peptide substrates. To gain further insight into the catalytic mechanism of PBP 5, we determined the three-dimensional structure of the G105D mutant form of soluble PBP 5 (termed sPBP 5') at 2.3 A resolution. The structure is composed of two domains, a penicillin binding domain with a striking similarity to Class A beta-lactamases (TEM-1-like) and a domain of unknown function. In addition, the penicillin-binding domain contains an active site loop spatially equivalent to the Omega loop of beta-lactamases. In beta-lactamases, the Omega loop contains two amino acids involved in catalyzing deacylation. This similarity may explain the high beta-lactamase activity of wild-type PBP 5. Because of the low rate of deacylation of the G105D mutant, visualization of peptide substrates bound to the active site may be possible. |
| Links |
PubMed Online version:10.1074/jbc.M004471200 |
| Keywords |
Acylation; Amino Acid Sequence; Bacterial Proteins; Binding Sites; Carrier Proteins/chemistry; Carrier Proteins/genetics; Carrier Proteins/metabolism; Crystallography, X-Ray; Escherichia coli/enzymology; Hexosyltransferases; Kinetics; Models, Molecular; Molecular Sequence Data; Muramoylpentapeptide Carboxypeptidase/chemistry; Muramoylpentapeptide Carboxypeptidase/genetics; Muramoylpentapeptide Carboxypeptidase/metabolism; Mutation/genetics; Penicillin-Binding Proteins; Penicillins/metabolism; Peptidyl Transferases; Protein Binding; Protein Structure, Secondary; Protein Structure, Tertiary; Sequence Alignment; beta-Lactamases/chemistry |
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