title: Redox regulation of OxyR requires specific disulfide bond formation involving a rapid kinetic reaction path.
abstract: The Escherichia coli OxyR transcription factor is activated by cellular hydrogen peroxide through the oxidation of reactive cysteines. Although there is substantial evidence for specific disulfide bond formation in the oxidative activation of OxyR, the presence of the disulfide bond has remained controversial. By mass spectrometry analyses and in vivo labeling assays we found that oxidation of OxyR in the formation of a specific disulfide bond between Cys199 and Cys208 in the wild-type protein. In addition, using time-resolved kinetic analyses, we determined that OxyR activation occurs at a rate of 9.7 s(-1). The disulfide bond-mediated conformation switch results in a metastable form that is locally strained by approximately 3 kcal mol(-1). On the basis of these observations we conclude that OxyR activation requires specific disulfide bond formation and that the rapid kinetic reaction path and conformation strain, respectively, drive the oxidation and reduction of OxyR.
authors.affiliation: Center for Cellular Switch Protein Structure, Korea Research Institute of Bioscience and Biotechnology, 52 Euh-eun-dong, Yuseong-gu, Daejeon 305-806, Korea.
authors.forename: Cheolju
authors.initials: C
authors.lastname: Lee
authors.forename: Soon Mi
authors.initials: SM
authors.lastname: Lee
authors.forename: Partha
authors.initials: P
authors.lastname: Mukhopadhyay
authors.forename: Seung Jun
authors.initials: SJ
authors.lastname: Kim
authors.forename: Sang Chul
authors.initials: SC
authors.lastname: Lee
authors.forename: Woo-Sung
authors.initials: WS
authors.lastname: Ahn
authors.forename: Myeong-Hee
authors.initials: MH
authors.lastname: Yu
authors.forename: Gisela
authors.initials: G
authors.lastname: Storz
authors.forename: Seong Eon
authors.initials: SE
authors.lastname: Ryu
mesh_terms: D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004220:Disulfides / Q000737:chemistry / Q000378:metabolism*; D004926:Escherichia coli / Q000737:chemistry / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins; D007700:Kinetics; D008958:Models, Molecular; D016877:Oxidants / Q000494:pharmacology; D010084:Oxidation-Reduction / Q000187:drug effects; D011489:Protein Denaturation / Q000187:drug effects; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D012097:Repressor Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013816:Thermodynamics; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D014364:Tryptophan / Q000378:metabolism; D014508:Urea / Q000494:pharmacology
chemical_list: D004268:DNA-Binding Proteins; D004220:Disulfides; D029968:Escherichia coli Proteins; D016877:Oxidants; D012097:Repressor Proteins; D014157:Transcription Factors; C499299:oxyR protein, E coli; D014364:Tryptophan; D014508:Urea