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Redox Partners (redox + partner)
Selected AbstractsCloning, purification, crystallization and preliminary X-ray analysis of a chimeric NADPH-cytochrome P450 reductaseACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 3 2009Louise Aigrain NADPH-cytochrome P450 reductase (CPR) is the favoured redox partner of microsomal cytochromes P450. This protein is composed of two flavin-containing domains (FMN and FAD) connected by a structured linker. An active CPR chimera consisting of the yeast FMN and human FAD domains has been produced, purified and crystallized. The crystals belonged to the monoclinic space group C2 and contained one molecule per asymmetric unit. Molecular replacement was performed using the published rat and yeast structures as search models. The initial electron-density maps revealed that the chimeric enzyme had crystallized in a conformation that differed from those of previously solved structures. [source] Redox-dependent structural changes in archaeal and bacterial Rieske-type [2Fe-2S] clustersPROTEIN SCIENCE, Issue 12 2002Nathaniel J. Cosper Abstract Proteins containing Rieske-type [2Fe-2S] clusters play important roles in many biological electron transfer reactions. Typically, [2Fe-2S] clusters are not directly involved in the catalytic transformation of substrate, but rather supply electrons to the active site. We report herein X-ray absorption spectroscopic (XAS) data that directly demonstrate an average increase in the iron,histidine bond length of at least 0.1 Å upon reduction of two distantly related Rieske-type clusters in archaeal Rieske ferredoxin from Sulfolobus solfataricus strain P-1 and bacterial anthranilate dioxygenases from Acinetobacter sp. strain ADP1. This localized redox-dependent structural change may fine tune the protein,protein interaction (in the case of ARF) or the interdomain interaction (in AntDO) to facilitate rapid electron transfer between a lower potential Rieske-type cluster and its redox partners, thereby regulating overall oxygenase reactions in the cells. [source] Structure of the M148Q mutant of rusticyanin at 1.5,Å: a model for the copper site of stellacyaninACTA CRYSTALLOGRAPHICA SECTION D, Issue 3 2001Michael A. Hough The small blue copper protein rusticyanin from Thiobacillus ferrooxidans contains a type 1 Cu centre with a single axial ligand, Met148, which together with the His-Cys-His trigonal planar ligands produces a distorted trigonal pyramidal coordination geometry to copper. Type 1 Cu sites are found in cupredoxins and several multicopper proteins, including oxidases and nitrite reductases. The role of the axial ligand has been extensively debated in terms of its function in the fine tuning of the redox potential and spectroscopic properties of type 1 Cu sites. Numerous mutations of the Met ligand in azurins have been studied, but interpretation of the results has been complicated by the presence of the additional carbonyl oxygen ligand from Gly45, a neighbouring residue to the coordinating His46. The importance of the axial ligand has been further emphasized by the finding that the type 1 centre in Rhus vernicifera stellacyanin, with the lowest redox potential in a type 1 Cu site of 184,mV, has Gln as the axial ligand, whilst fungal laccase and ceruloplasmin, which have redox potentials of 550,800,mV, have a Leu in this position. Here, the crystal structure of the M148Q mutant of rusticyanin at 1.5,Å resolution is presented. This is a significantly higher resolution than that of the structures of native rusticyanin. In addition, the M148Q structure is that of the oxidized protein while the native structures to date are of the reduced protein. The mutant protein crystallizes with two molecules per asymmetric unit, in contrast to the one present in the native crystal form. This mutant's redox potential (550,mV at pH 3.2) is lowered compared with that of the native protein (,670,mV at pH 3.2) by about 120,mV. The type 1 Cu site of M148Q closely mimics the structural characteristics of the equivalent site in non-glycosylated cucumber stellacyanin (redox potential ,260,mV) and, owing to the absence in rusticyanin of the fifth, carbonyl ligand present in azurin, may provide a better model for the R. vernicifera stellacyanin (redox potential ,184,mV) type 1 Cu site, which also lacks the fifth ligand. Furthermore, the presence of two molecules in the asymmetric unit cell indicates a potential binding region of the redox partners. [source] Discovery of 2-Naphthoic Acid Monooxygenases by Genome Mining and their Use as BiocatalystsCHEMSUSCHEM CHEMISTRY AND SUSTAINABILITY, ENERGY & MATERIALS, Issue 7 2009Toshiki Furuya Dr. Abstract The large pool of cytochrome P450 (P450) open-reading frames identified in genome sequences has attracted much attention as a resource for new oxidation biocatalysts. P450 genes were cloned from genome-sequenced bacteria and coexpressed with putidaredoxin and its reductase genes to provide the redox partners of P450 in Escherichia coli. Whole-cell assays were performed with 2-naphthoic acid as a substrate. Hydroxylated naphthoic acid products were rapidly detected with two reagents showing different colors in the presence of the products. Two P450s, CYP199A1 and CYP199A2, were found to hydroxylate the substrate to 7- and 8-hydroxy-2-naphthoic acids. The CYP199A1 whole-cell biocatalyst converted 1,mM 2-naphthoic acid to 0.27,mM 7-hydroxy-2-naphthoic acid and 0.53,mM 8-hydroxy-2-naphthoic acid. CYP199A2 exhibited similar regioselectivity to CYP199A1. Furthermore, we found that 8-hydroxy-2-naphthoic acid emits near-white fluorescence when exposed to UV light. These P450s will provide a facile and environmentally friendly synthetic approach to the hydroxynaphthoic acids. [source] | ||||||||||