Isolation of partly heme devoid but not fully carbon monoxide reacting cytochrome b, from beef heart mitochondria.
An easy and rapid preparation of cytochrome b from beef heart mitochondria is described. The method is based on four fractionation steps in the presence of bile acid and one step DE chromatography. The isolated cytochrome shows only partial reactivity with carbon monoxide; up to a level of 35 micrograms protein applied, it moves as a single band on polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, with an apparent molecular weight of 32000 +/- 1000. A ratio of 14.1 +/- 0.8 nmol heme b/mg protein has been found for isolated cytochrome, indicating that good part of heme b is lost during the isolation. The low number of steps required for the isolation could allow the identification of the stage(s) responsible for the loss of heme moiety and indicate preparation improvements.
D000818:Animals; D002248:Carbon Monoxide / Q000378:metabolism*; D002417:Cattle; D003573:Cytochrome b Group / Q000302:isolation & purification*; D006418:Heme / Q000032:analysis*; D008722:Methods; D008929:Mitochondria, Heart / Q000032:analysis*; D013053:Spectrophotometry
D003573:Cytochrome b Group; D006418:Heme; D002248:Carbon Monoxide