Purification and Characterization of Microsomal Cytochrome b5 and NADH Cytochrome b5 Reductase from Pisum sativum | Zendy

David R. Jollie | Zendy; Stephen G. Sligar | Zendy; Mary A. Schuler | Zendy

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Purification and Characterization of Microsomal Cytochrome b₅ and NADH Cytochrome b₅ Reductase from Pisum sativum

Author(s) -

David R. Jollie,

Stephen G. Sligar,

Mary A. Schuler

Publication year - 1987

Publication title -

plant physiology

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 3.554

H-Index - 312

eISSN - 1532-2548

pISSN - 0032-0889

DOI - 10.1104/pp.85.2.457

Subject(s) - biochemistry , cytochrome , pisum , reductase , cytochrome b , microsome , cytochrome c , cytochrome c oxidase , biology , cytochrome p450 reductase , oxidase test , amino acid , chemistry , enzyme , coenzyme q – cytochrome c reductase , mitochondrion , gene , mitochondrial dna

In this communication we document the reproducible protocols for the purification of milligram quantities of cytochrome b(5) and NADH-cytochrome b(5) reductase from the microsomal fraction of Pisum sativum. The cytochrome b(5) component of this NADH linked electron transport chain was found to have a molecular mass of 16,400 daltons and the reductase a molecular mass of 34,500 daltons. These components could be reconstituted into a functional NADH oxidase activity active in the reduction of exogenous cytochrome c or ferricyanide. In the latter assay the purified reductase exhibited a turnover number of 22,000 per minute. The amino-terminal amino acid sequence of the cytochrome b(5) component was determined by sequential Edmund degredation, thus providing crucial information for the efficient cloning of this central protein of plant microsomal electron transfer.

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