title: Impact of mutations in DNA gyrase genes on quinolone resistance in Campylobacter jejuni.
abstract: Amino acid substitutions providing quinolone resistance to Campyloabcter jejuni have been found in the quinolone resistance-determining region of protein DNA gyrase subunit A (GyrA), with the highest frequency at position 86 followed by position 90. In this study, wild-type and mutant recombinant DNA gyrase subunits were expressed in Escherichia coli and purified using Ni-NTA agarose column chromatography. Soluble 97 kDa GyrA and 87 kDa DNA gyrase subunit B were shown to reconstitute ATP-dependent DNA supercoiling activity. A quinolone-inhibited supercoiling assay demonstrated the roles of Thr86Ile, Thr86Ala, Thr86Lys, Asp90Asn, and Asp90Tyr amino acid substitutions in reducing sensitivity to quinolones. The marked effect of Thr86Ile on all examined quinolones suggested the advantage of this substitution in concordance with recurring isolation of quinolone-resistant C. jejuni. An analysis of the structure-activity relationship showed the importance of the substituent at position 8 in quinolones to overcome the effect of Thr86Ile. Sitafloxacin (SIT), which has a fluorinate cyclopropyl ring at R-1 and a chloride substituent at R-8, a characteristic not found in other quinolones, showed the highest inhibitory activity against all mutant C. jejuni gyrases including ciprofloxacin-resistant mutants. The results suggest SIT as a promising drug for the treatment of campylobacteriosis caused by CIP-resistant C. jejuni. Copyright © 2016 John Wiley & Sons, Ltd.
authors: Division of Bioresources, Hokkaido University Research Center for Zoonosis Control, Sapporo, Japan.
authors: Ruchirada
authors: R
authors: Changkwanyeun
authors: Division of Bioresources, Hokkaido University Research Center for Zoonosis Control, Sapporo, Japan.
authors: Tomoyuki
authors: T
authors: Yamaguchi
authors: Division of Bioresources, Hokkaido University Research Center for Zoonosis Control, Sapporo, Japan.
authors: Siriporn
authors: S
authors: Kongsoi
authors: Division of Bioresources, Hokkaido University Research Center for Zoonosis Control, Sapporo, Japan.
authors: Kanjana
authors: K
authors: Changkaew
authors: Central Research Laboratory, Kissei Pharmaceutical Co., Ltd, Nagano, Japan.
authors: Kazumasa
authors: K
authors: Yokoyama
authors: Laboratory of Tuberculosis, Department of Bacteriology II, National Institute Infectious Diseases, Tokyo, Japan.
authors: Hyun
authors: H
authors: Kim
authors: Faculty of Public Health, Thammasat University, Rangsit, Thailand.
authors: Orasa
authors: O
authors: Suthienkul
authors: School of Veterinary Medicine, Rakuno Gakuen University, Ebetsu, Hokkaido, Japan.
authors: Masaru
authors: M
authors: Usui
authors: School of Veterinary Medicine, Rakuno Gakuen University, Ebetsu, Hokkaido, Japan.
authors: Yutaka
authors: Y
authors: Tamura
authors: Division of Bioresources, Hokkaido University Research Center for Zoonosis Control, Sapporo, Japan.|Hokkaido University The Global station for Zoonosis Control, Sapporo, Japan.
authors: Chie
authors: C
authors: Nakajima
authors: Division of Bioresources, Hokkaido University Research Center for Zoonosis Control, Sapporo, Japan. suzuki@czc.hokudai.ac.jp.|Hokkaido University The Global station for Zoonosis Control, Sapporo, Japan. suzuki@czc.hokudai.ac.jp.
authors: Yasuhiko
authors: Y
authors: Suzuki
mesh_terms: D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000494:pharmacology*; D016123:Campylobacter jejuni / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D002939:Ciprofloxacin / Q000737:chemistry / Q000494:pharmacology*; D027081:DNA Gyrase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000737:chemistry* / Q000378:metabolism; D024841:Fluoroquinolones / Q000737:chemistry / Q000494:pharmacology*; D008826:Microbial Sensitivity Tests; D009154:Mutation / Q000187:drug effects*; D015363:Quinolones / Q000737:chemistry / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000737:chemistry* / Q000378:metabolism; D013329:Structure-Activity Relationship; D059005:Topoisomerase II Inhibitors / Q000494:pharmacology*
chemical_list: D000900:Anti-Bacterial Agents; D024841:Fluoroquinolones; D015363:Quinolones; D011994:Recombinant Proteins; D059005:Topoisomerase II Inhibitors; D002939:Ciprofloxacin; C076246:sitafloxacin; D027081:DNA Gyrase
keywords: Campylobacter jejuni; DNA gyrase; mutation; quinolone resistance