title: Site-directed mutagenesis and chemical modification of the two cysteine residues of the UDP-N-acetylmuramoyl:L-alanine ligase of Escherichia coli.
abstract: Site-directed mutagenesis and chemical modification of the two cysteine residues of the MurC L-alanine-adding enzyme from Escherichia coli were undertaken to study their possible role in activity and stability. Their replacement by alanine was not critical for activity. However, C230 played a role in enzyme stability and substrate binding. N-Ethylmaleimide alkylation led to monoalkylated and dialkylated proteins. The monoalkylated protein had mostly unmodified C230 residues. The extent of alkylation of C230 paralleled the loss of activity, whereas that of C426 did not. Protection against inactivation by beta,gamma-imidoadenosine 5'-triphosphate implied the involvement of C230 in the ATP binding site.
authors.affiliation: Biophysics, Hoechst-Marion-Roussel, Romainville, France.
authors.forename: F
authors.initials: F
authors.lastname: Nosal
authors.forename: A
authors.initials: A
authors.lastname: Masson
authors.forename: R
authors.initials: R
authors.lastname: Legrand
authors.forename: D
authors.initials: D
authors.lastname: Blanot
authors.forename: B
authors.initials: B
authors.lastname: Schoot
authors.forename: J
authors.initials: J
authors.lastname: van Heijenoort
authors.forename: C
authors.initials: C
authors.lastname: Parquet
mesh_terms: D000409:Alanine / Q000235:genetics; D000478:Alkylation; D003545:Cysteine / Q000235:genetics*; D004789:Enzyme Activation / Q000187:drug effects / Q000235:genetics; D004795:Enzyme Stability / Q000187:drug effects / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics*; D005033:Ethylmaleimide / Q000494:pharmacology; D016297:Mutagenesis, Site-Directed; D010449:Peptide Mapping; D010453:Peptide Synthases / Q000737:chemistry / Q000235:genetics*; D019163:Reducing Agents / Q000494:pharmacology
chemical_list: D019163:Reducing Agents; D010453:Peptide Synthases; C023229:UDP-N-acetylmuramoyl-alanine synthetase; D003545:Cysteine; D005033:Ethylmaleimide; D000409:Alanine