title: Mutation of the gene encoding monothiol glutaredoxin (GrxD) in Pseudomonas aeruginosa increases its susceptibility to polymyxins.
abstract: Pseudomonas aeruginosa is a frequent cause of hospital-acquired infections that have a high mortality rate because of its innate drug resistance. Polymyxins are recognised as the last-line antibiotics for the treatment of multidrug-resistant (MDR) P. aeruginosa. In this study, the link between monothiol glutaredoxin (GrxD), which catalyses the reduction of disulphide bonds of various substrates in P. aeruginosa, and antibiotic resistance was examined. A P. aeruginosa ΔgrxD mutant strain was constructed. The ΔgrxD mutant showed significantly increased susceptibility to polymyxin B (PMB) compared with the wild-type P. aeruginosa PAO1. Site-directed mutagenesis was performed to generate amino acid substitutions in GrxD, and the ability of mutated grxD genes to confer resistance to PMB in the ΔgrxD mutant was tested. The results indicated that residue C29 at the active site of GrxD is important for protection against polymyxin killing in the mutant. Polymyxin killing of PAO1 and the ΔgrxD mutant did not appear to involve hydroxyl radicals generated by antibiotic treatment because increased susceptibility of the mutant to PMB was also observed under anaerobic growth as well as aerobically in the presence of the iron chelator 2,2'-dipyridyl. Thus, GrxD could be a target for the development of agents that enhance the effectiveness of PMB in treating clinically important MDR P. aeruginosa infections.
authors.affiliation: Laboratory of Biotechnology, Chulabhorn Research Institute, Bangkok 10210, Thailand.
authors.forename: Adisak
authors.initials: A
authors.lastname: Romsang
authors.affiliation: Molecular Medicine Graduate Program, Faculty of Science, Mahidol University, Bangkok 10400, Thailand.
authors.forename: Panithi
authors.initials: P
authors.lastname: Leesukon
authors.affiliation: Laboratory of Biotechnology, Chulabhorn Research Institute, Bangkok 10210, Thailand.
authors.forename: Jintana
authors.initials: J
authors.lastname: Duangnkern
authors.affiliation: Laboratory of Biotechnology, Chulabhorn Research Institute, Bangkok 10210, Thailand; Center of Excellence on Environmental Health and Toxicology, Ministry of Education, Bangkok, Thailand.
authors.forename: Paiboon
authors.initials: P
authors.lastname: Vattanaviboon
authors.affiliation: Laboratory of Biotechnology, Chulabhorn Research Institute, Bangkok 10210, Thailand; Center of Excellence on Environmental Health and Toxicology, Ministry of Education, Bangkok, Thailand; Center for Emerging Bacterial Infections, Faculty of Science, Mahidol University, Bangkok 10400, Thailand; Department of Biotechnology, Faculty of Science, Mahidol University, Bangkok 10400, Thailand. Electronic address: skorn@cri.or.th.
authors.forename: Skorn
authors.initials: S
authors.lastname: Mongkolsuk
mesh_terms: D019943:Amino Acid Substitution; D004252:DNA Mutational Analysis; D024881:Drug Resistance, Bacterial; D017353:Gene Deletion; D054477:Glutaredoxins / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D008826:Microbial Sensitivity Tests; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D020125:Mutation, Missense; D011113:Polymyxins / Q000494:pharmacology*; D011550:Pseudomonas aeruginosa / Q000187:drug effects* / Q000201:enzymology*
chemical_list: D054477:Glutaredoxins; D050505:Mutant Proteins; D011113:Polymyxins
keywords: Monothiol glutaredoxin; Polymyxins; Pseudomonas aeruginosa