title: Point mutations of two arginine residues in the Streptomyces R61 DD-peptidase.
abstract: Incubation of the exocellular DD-carboxypeptidase/transpeptidase of Streptomyces R61 with phenylglyoxal resulted in a time-dependent decrease in the enzyme activity. This inactivation was demonstrated to be due to modification of the Arg-99 side chain. In consequence, the role of that residue was investigated by site-directed mutagenesis. Mutation of Arg-99 into leucine appeared to be highly detrimental to enzyme stability, reflecting a determining structural role for this residue. The conserved Arg-103 residue was also substituted by using site-directed mutagenesis. The modification to a serine residue yielded a stable enzyme, the catalytic properties of which were similar to those of the wild-type enzyme. Thus Arg-103, although strictly conserved or replaced by a lysine residue in most of the active-site penicillin-recognizing proteins, did not appear to fulfil any essential role in either the enzyme activity or structure.
authors.affiliation: Laboratoire d'Enzymologie, Université de Liège, Sart-Tilman, Belgium.
authors.forename: C
authors.initials: C
authors.lastname: Bourguignon-Bellefroid
authors.forename: B
authors.initials: B
authors.lastname: Joris
authors.forename: J
authors.initials: J
authors.lastname: Van Beeumen
authors.forename: J M
authors.initials: JM
authors.lastname: Ghuysen
authors.forename: J M
authors.initials: JM
authors.lastname: Frère
mesh_terms: D000595:Amino Acid Sequence; D001120:Arginine / Q000235:genetics*; D001426:Bacterial Proteins / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D004789:Enzyme Activation; D007930:Leucine / Q000235:genetics; D008969:Molecular Sequence Data; D002267:Muramoylpentapeptide Carboxypeptidase / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D009154:Mutation; D010406:Penicillins / Q000378:metabolism; D010658:Phenylglyoxal / Q000494:pharmacology; D011485:Protein Binding; D012694:Serine / Q000235:genetics; D013302:Streptomyces / Q000201:enzymology* / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D010406:Penicillins; D012694:Serine; D001120:Arginine; D002267:Muramoylpentapeptide Carboxypeptidase; D007930:Leucine; D010658:Phenylglyoxal