title: Purification and partial characterization of microsomal NADH-cytochrome b5 reductase from higher plant Catharanthus roseus.
abstract: A simple three step procedure was used to purify microsomal NADH-cytochrome b5 (ferricyanide) reductase to homogeneity from the higher plant C. roseus. The microsomal bound reductase was solubilized using zwitterionic detergent-CHAPS. The solubilized reductase was subjected to affinity chromatography on octylamino Sepharose 4B, blue 2-Sepharose CL-6B and NAD(+)-Agarose. The homogeneous enzyme has an apparent molecular weight of 33,000 as estimated by SDS-PAGE. The purified enzyme catalyzes the reduction of purified cytochrome b5 from C. roseus in the presence of NADH. The reductase also readily transfers electrons from NADH to ferricyanide (Km 56 microM), 2, 6-dichlorophenolindophenol (Km 65 microM) and cytochrome c via cytochrome b5 but not to menadione.
authors.affiliation: Department of Organic Chemistry, Indian Institute of Science, Bangalore.
authors.forename: K M
authors.initials: KM
authors.lastname: Madyastha
authors.forename: N K
authors.initials: NK
authors.lastname: Chary
authors.forename: R
authors.initials: R
authors.lastname: Holla
authors.forename: T B
authors.initials: TB
authors.lastname: Karegowdar
mesh_terms: D002846:Chromatography, Affinity; D003579:Cytochrome Reductases / Q000302:isolation & purification* / Q000378:metabolism*; D042966:Cytochrome-B(5) Reductase; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D008861:Microsomes / Q000201:enzymology*; D008970:Molecular Weight; D010944:Plants / Q000201:enzymology*
chemical_list: D003579:Cytochrome Reductases; D042966:Cytochrome-B(5) Reductase