title: Key roles of the Escherichia coli AhpC C-terminus in assembly and catalysis of alkylhydroperoxide reductase, an enzyme essential for the alleviation of oxidative stress.
abstract: 2-Cys peroxiredoxins (Prxs) are a large family of peroxidases, responsible for antioxidant function and regulation in cell signaling, apoptosis and differentiation. The Escherichia coli alkylhydroperoxide reductase (AhpR) is a prototype of the Prxs-family, and is composed of an NADH-dependent AhpF reductase (57 kDa) and AhpC (21 kDa), catalyzing the reduction of H2O2. We show that the E. coli AhpC (EcAhpC, 187 residues) forms a decameric ring structure under reduced and close to physiological conditions, composed of five catalytic dimers. Single particle analysis of cryo-electron micrographs of C-terminal truncated (EcAhpC1 -172 and EcAhpC1 -182) and mutated forms of EcAhpC reveals the loss of decamer formation, indicating the importance of the very C-terminus of AhpC in dimer to decamer transition. The crystallographic structures of the truncated EcAhpC1 -172 and EcAhpC1 -182 demonstrate for the first time that, in contrast to the reduced form, the very C-terminus of the oxidized EcAhpC is oriented away from the AhpC dimer interface and away from the catalytic redox-center, reflecting structural rearrangements during redox-modulation and -oligomerization. Furthermore, using an ensemble of different truncated and mutated EcAhpC protein constructs the importance of the very C-terminus in AhpC activity and in AhpC-AhpF assembly has been demonstrated.
authors.affiliation: Nanyang Technological University, School of Biological Sciences, 60 Nanyang Drive, Singapore 637551; Program in Emerging Infectious Diseases, Duke-NUS Graduate Medical School, KTP Building, 8 College Road, Singapore 169857; Center for Bioimaging Sciences, Dept. of Biological Sciences, National University of Singapore, Singapore 119077.
authors.forename: Phat Vinh
authors.initials: PV
authors.lastname: Dip
authors.affiliation: Bioinformatics Institute, Agency for Science, Technology and Research (A*STAR), 30 Biopolis Street, #07-01 Matrix, Singapore 138671.
authors.forename: Neelagandan
authors.initials: N
authors.lastname: Kamariah
authors.affiliation: Nanyang Technological University, School of Biological Sciences, 60 Nanyang Drive, Singapore 637551.
authors.forename: Wilson
authors.initials: W
authors.lastname: Nartey
authors.affiliation: Nanyang Technological University, School of Biological Sciences, 60 Nanyang Drive, Singapore 637551.
authors.forename: Claudia
authors.initials: C
authors.lastname: Beushausen
authors.affiliation: Program in Emerging Infectious Diseases, Duke-NUS Graduate Medical School, KTP Building, 8 College Road, Singapore 169857; Center for Bioimaging Sciences, Dept. of Biological Sciences, National University of Singapore, Singapore 119077.
authors.forename: Victor A
authors.initials: VA
authors.lastname: Kostyuchenko
authors.affiliation: Program in Emerging Infectious Diseases, Duke-NUS Graduate Medical School, KTP Building, 8 College Road, Singapore 169857; Center for Bioimaging Sciences, Dept. of Biological Sciences, National University of Singapore, Singapore 119077.
authors.forename: Thiam-Seng
authors.initials: TS
authors.lastname: Ng
authors.affiliation: Program in Emerging Infectious Diseases, Duke-NUS Graduate Medical School, KTP Building, 8 College Road, Singapore 169857; Center for Bioimaging Sciences, Dept. of Biological Sciences, National University of Singapore, Singapore 119077.
authors.forename: Shee-Mei
authors.initials: SM
authors.lastname: Lok
authors.affiliation: Nanyang Technological University, School of Biological Sciences, 60 Nanyang Drive, Singapore 637551.
authors.forename: Wuan Geok
authors.initials: WG
authors.lastname: Saw
authors.affiliation: Bioinformatics Institute, Agency for Science, Technology and Research (A*STAR), 30 Biopolis Street, #07-01 Matrix, Singapore 138671; School of Computer Engineering, Nanyang Technological University, 50 Nanyang Drive, Singapore 637553; Department of Biological Sciences, National University of Singapore, 8 Medical Drive, Singapore 117597.
authors.forename: Frank
authors.initials: F
authors.lastname: Eisenhaber
authors.affiliation: Bioinformatics Institute, Agency for Science, Technology and Research (A*STAR), 30 Biopolis Street, #07-01 Matrix, Singapore 138671.
authors.forename: Birgit
authors.initials: B
authors.lastname: Eisenhaber
authors.affiliation: Nanyang Technological University, School of Biological Sciences, 60 Nanyang Drive, Singapore 637551; Bioinformatics Institute, Agency for Science, Technology and Research (A*STAR), 30 Biopolis Street, #07-01 Matrix, Singapore 138671. Electronic address: ggrueber@ntu.edu.sg.
authors.forename: Gerhard
authors.initials: G
authors.lastname: Grüber
mesh_terms: D055162:Biocatalysis; D020285:Cryoelectron Microscopy; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D009154:Mutation; D009243:NAD / Q000737:chemistry / Q000378:metabolism; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D054464:Peroxiredoxins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D055503:Protein Multimerization; D013053:Spectrophotometry
chemical_list: D029968:Escherichia coli Proteins; D009243:NAD; D054464:Peroxiredoxins; C515937:ahpC protein, E coli; C515938:ahpF protein, E coli