ISOLATION AND CHARACTERIZATION OF CYTOCHROME b-559 FROM CHLOROPLASTS AND ETIOPLASTS OF BARLEY

A procedure permitting the isolation of large amounts of cytochrome b-559 from chloroplasts of barley is described. The isolated cytochrome b-559 has an apparent molecular weight of 120,000. The protein equivalent weight per mole of heme is 30,000. Depending on the electrophoretic conditions used, this molecule can be dissociated into components with apparent molecular weights of 31,000, 30,000, 18,000 and 9,000. The two polypeptides at M r 31,000 and 30,000 cannot be stained with Coomassie Brilliant Blue R250 but are stained with tetramethyl benzidine, a reagent which can be used to detect heme-associated peroxidase activity. An isolation procedure using less denaturing conditions resulted in the isolation of cytochrome b-559 together with polypeptides of Mr 33,000, 21,000 and 18,000 all of which are stained with Coomassie Brilliant Blue. The low potential form of cytochrome b-559 present in etioplasts was shown to be extremely labile compared to the low potential form of cytochrome b-559 isolated from chloroplasts. However, an antibody raised against chloroplast cytochrome b-559 also reacted with the form isolated from etioplasts and agglutinated chloroplast as well as etioplast lamellae.