title: Generation, characterization and crystallization of a cytochrome c(1)-subunit IV fused cytochrome bc(1) complex from Rhodobacter sphaeroides.
abstract: Cytochrome bc(1) complex catalyzes the reaction of electron transfer from ubiquinol to cytochrome c (or cytochrome c(2)) and couples this reaction to proton translocation across the membrane. Crystallization of the Rhodobacter sphaeroides bc(1) complex resulted in crystals containing only three core subunits. To mitigate the problem of subunit IV being dissociated from the three-subunit core complex during crystallization, we recently engineered an R. sphaeroides mutant in which the N-terminus of subunit IV was fused to the C-terminus of cytochrome c(1) with a 14-glycine linker between the two fusing subunits, and a 6-histidine tag at the C-terminus of subunit IV (c(1)-14Gly-IV-6His). The purified fusion mutant complex shows higher electron transfer activity, more structural stability, and less superoxide generation as compared to the wild-type enzyme. Preliminary crystallization attempts with this mutant complex yielded crystals containing four subunits and diffracting X-rays to 5.5Å resolution.
authors: Department of Biochemistry & Molecular Biology, Oklahoma State University, Stillwater, OK 74078, USA.
authors: Ting
authors: T
authors: Su
authors: Lothar
authors: L
authors: Esser
authors: Di
authors: D
authors: Xia
authors: Chang-An
authors: CA
authors: Yu
authors: Linda
authors: L
authors: Yu
mesh_terms: D000818:Animals; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D003575:Cytochromes c1 / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D014450:Electron Transport Complex III / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D006736:Horses; D008958:Models, Molecular; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D030781:Organisms, Genetically Modified; D010788:Photosynthesis / Q000235:genetics; D017433:Protein Structure, Secondary; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D012242:Rhodobacter sphaeroides / Q000235:genetics* / Q000378:metabolism
chemical_list: D050505:Mutant Proteins; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins; D003575:Cytochromes c1; D014450:Electron Transport Complex III