title: Engineered Citrobacter freundii methionine γ-lyase effectively produces antimicrobial thiosulfinates.
abstract: Antimicrobial activity of thiosulfinates in situ produced by mixtures of Citrobacter freundii methionine γ-lyase (MGL) with new substrates, l-methionine and S-(alkyl/allyl)-l-cysteine sulfoxides has been recently demonstrated (Anufrieva et al., 2015). This opens a way to the rational design of a new biotechnologically relevant antimicrobial drug producer. To increase the efficiency of the enzyme toward sulfoxides, the mutant forms of MGL, with the replacements of active site cysteine 115 with alanine (C115A MGL) and histidine (C115H MGL) were obtained. The replacement of cysteine 115 by histidine results in the loss of activity of the mutant enzyme in the γ-elimination reaction of physiological substrate, whereas the activity in the β-elimination reaction of characteristic substrates persists. However, the catalytic efficiency of C115H MGL in the β-elimination reaction of S-substituted l-cysteine sulfoxides is increased by about an order of magnitude compared to the wild type MGL. The antibacterial activity of C115H MGL mixtures with a number of sulfoxides was assessed against Gram-positive and Gram-negative bacteria. The bacteriostatic effect was more pronounced against Gram-positive than against Gram-negative bacteria, while antibacterial potential proved to be quite similar. Thus, the mutant enzyme C115H MGL is an effective catalyst, in particular, for decomposition of sulfoxides and the pharmacological couples of the mutant form with sulfoxides might be new antimicrobial agents.
authors.affiliation: Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilova Str., 32, Moscow, 119991, Russia.
authors.forename: Elena A
authors.initials: EA
authors.lastname: Morozova
authors.affiliation: Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilova Str., 32, Moscow, 119991, Russia.
authors.forename: Vitalia V
authors.initials: VV
authors.lastname: Kulikova
authors.affiliation: Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilova Str., 32, Moscow, 119991, Russia.
authors.forename: Alexei N
authors.initials: AN
authors.lastname: Rodionov
authors.affiliation: Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilova Str., 32, Moscow, 119991, Russia.
authors.forename: Svetlana V
authors.initials: SV
authors.lastname: Revtovich
authors.affiliation: Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilova Str., 32, Moscow, 119991, Russia.
authors.forename: Natalya V
authors.initials: NV
authors.lastname: Anufrieva
authors.affiliation: Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilova Str., 32, Moscow, 119991, Russia. Electronic address: tvd@eimb.ru.
authors.forename: Tatyana V
authors.initials: TV
authors.lastname: Demidkina
mesh_terms: D000409:Alanine / Q000235:genetics / Q000378:metabolism; D000890:Anti-Infective Agents / Q000378:metabolism* / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D055162:Biocatalysis; D013437:Carbon-Sulfur Lyases / Q000235:genetics / Q000378:metabolism*; D016971:Citrobacter freundii / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D006639:Histidine / Q000235:genetics / Q000378:metabolism; D060847:Metabolic Engineering / Q000379:methods; D008715:Methionine / Q000378:metabolism; D008826:Microbial Sensitivity Tests; D020125:Mutation, Missense; D013053:Spectrophotometry; D013379:Substrate Specificity; D013441:Sulfinic Acids / Q000378:metabolism* / Q000494:pharmacology; D013454:Sulfoxides / Q000378:metabolism
chemical_list: D000890:Anti-Infective Agents; D001426:Bacterial Proteins; D013441:Sulfinic Acids; D013454:Sulfoxides; C100040:thiosulfinic acids; D006639:Histidine; D008715:Methionine; D013437:Carbon-Sulfur Lyases; C021558:L-methionine gamma-lyase; D003545:Cysteine; D000409:Alanine
keywords: Antibacterial activity; Citrobacter freundii; Cysteine sulfoxides; Design of pharmacological couple; Methionine γ-lyase; Site-directed mutagenesis; Thiosulfinates