title: Structures of protein-protein complexes involved in electron transfer.
abstract: Electron transfer reactions are essential for life because they underpin oxidative phosphorylation and photosynthesis, processes leading to the generation of ATP, and are involved in many reactions of intermediary metabolism. Key to these roles is the formation of transient inter-protein electron transfer complexes. The structural basis for the control of specificity between partner proteins is lacking because these weak transient complexes have remained largely intractable for crystallographic studies. Inter-protein electron transfer processes are central to all of the key steps of denitrification, an alternative form of respiration in which bacteria reduce nitrate or nitrite to N2 through the gaseous intermediates nitric oxide (NO) and nitrous oxide (N2O) when oxygen concentrations are limiting. The one-electron reduction of nitrite to NO, a precursor to N2O, is performed by either a haem- or copper-containing nitrite reductase (CuNiR) where they receive an electron from redox partner proteins a cupredoxin or a c-type cytochrome. Here we report the structures of the newly characterized three-domain haem-c-Cu nitrite reductase from Ralstonia pickettii (RpNiR) at 1.01 Å resolution and its M92A and P93A mutants. Very high resolution provides the first view of the atomic detail of the interface between the core trimeric cupredoxin structure of CuNiR and the tethered cytochrome c domain that allows the enzyme to function as an effective self-electron transfer system where the donor and acceptor proteins are fused together by genomic acquisition for functional advantage. Comparison of RpNiR with the binary complex of a CuNiR with a donor protein, AxNiR-cytc551 (ref. 6), and mutagenesis studies provide direct evidence for the importance of a hydrogen-bonded water at the interface in electron transfer. The structure also provides an explanation for the preferential binding of nitrite to the reduced copper ion at the active site in RpNiR, in contrast to other CuNiRs where reductive inactivation occurs, preventing substrate binding.
authors.affiliation: Molecular Biophysics Group, Institute of Integrative Biology, Faculty of Health and Life Sciences, University of Liverpool, Liverpool L69 7ZX, UK.
authors.forename: Svetlana V
authors.initials: SV
authors.lastname: Antonyuk
authors.forename: Cong
authors.initials: C
authors.lastname: Han
authors.forename: Robert R
authors.initials: RR
authors.lastname: Eady
authors.forename: S Samar
authors.initials: SS
authors.lastname: Hasnain
mesh_terms: D001400:Azurin / Q000737:chemistry / Q000378:metabolism; D020134:Catalytic Domain; D003300:Copper / Q000737:chemistry / Q000378:metabolism; D045304:Cytochromes c / Q000737:chemistry / Q000378:metabolism; D004579:Electron Transport; D006860:Hydrogen Bonding; D008958:Models, Molecular; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009572:Nitrite Reductases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D009573:Nitrites / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011522:Protons; D049920:Ralstonia pickettii / Q000201:enzymology*; D014867:Water / Q000737:chemistry / Q000378:metabolism
chemical_list: D050505:Mutant Proteins; D009573:Nitrites; D011522:Protons; C050107:cupredoxin; D014867:Water; D001400:Azurin; D003300:Copper; D045304:Cytochromes c; D009572:Nitrite Reductases; C052695:nitrite reductase, copper-containing
references.citation: Acta Crystallogr D Biol Crystallogr. 1994 Sep 1;50(Pt 5):760-3
references.pmid: 15299374
references.citation: J Biol Inorg Chem. 2007 Nov;12(8):1119-27
references.pmid: 17712582
references.citation: Nat Struct Biol. 1995 Nov;2(11):975-82
references.pmid: 7583671
references.citation: J Mol Biol. 2003 Apr 25;328(2):429-38
references.pmid: 12691751
references.citation: Nature. 1992 Feb 27;355(6363):796-802
references.pmid: 1311417
references.citation: Dalton Trans. 2012 Mar 28;41(12):3355-68
references.pmid: 21918782
references.citation: Biochem J. 2012 Jun 1;444(2):219-26
references.pmid: 22414182
references.citation: J Am Chem Soc. 2007 Jul 11;129(27):8557-65
references.pmid: 17579406
references.citation: Nature. 2009 Nov 5;462(7269):117-20
references.pmid: 19890332
references.citation: J Biol Chem. 2000 Aug 4;275(31):23957-64
references.pmid: 10811642
references.citation: Biochemistry. 2004 Nov 9;43(44):14180-8
references.pmid: 15518568
references.citation: J Mol Biol. 1999 Apr 16;287(5):1001-9
references.pmid: 10222206
references.citation: Proc Natl Acad Sci U S A. 2005 Aug 23;102(34):12041-6
references.pmid: 16093314
references.citation: Proc Natl Acad Sci U S A. 2010 Jun 29;107(26):11799-804
references.pmid: 20547871
references.citation: Acta Crystallogr D Biol Crystallogr. 2010 Jan;66(Pt 1):12-21
references.pmid: 20057044
references.citation: Acta Crystallogr D Biol Crystallogr. 1997 May 1;53(Pt 3):240-55
references.pmid: 15299926
references.citation: Biochemistry. 2008 Dec 23;47(51):13547-53
references.pmid: 19053252
references.citation: Methods Enzymol. 1997;276:307-26
references.pmid: 27754618
references.citation: Microbiol Mol Biol Rev. 1997 Dec;61(4):533-616
references.pmid: 9409151
references.citation: Nature. 2012 Jul 26;487(7408):514-8
references.pmid: 22763450
references.citation: Chem Rev. 2006 Jan;106(1):90-115
references.pmid: 16402772
references.citation: Science. 2005 Nov 25;310(5752):1311-3
references.pmid: 16311331
references.citation: Proc Natl Acad Sci U S A. 2007 Mar 13;104(11):4315-20
references.pmid: 17360521
references.citation: Nat Struct Biol. 1994 Apr;1(4):234-8
references.pmid: 7656052
references.citation: Science. 1991 Jul 26;253(5018):438-42
references.pmid: 1862344
references.citation: Nat Struct Biol. 1995 Apr;2(4):287-92
references.pmid: 7796265
references.citation: Science. 2004 May 7;304(5672):867-70
references.pmid: 15131305
references.citation: Acta Crystallogr D Biol Crystallogr. 2004 Dec;60(Pt 12 Pt 1):2126-32
references.pmid: 15572765