Flavoproteins, absorption maxima

NADH-cytochrome P-450 reductase was purified as a single polypeptide chain with a molecular weight of 51 kDa. The absorption spectrum of the purified protein showed typical flavoprotein absorption maxima at 370 and 446 run, and a shoulder at 475 nm. Consistent with the detection of the flavin prosthetic group, the purified NADH-... 

Like the nicotinamide coenzymes (Fig. 13-15), the flavin nucleotides undergo a shift in a major absorption band on reduction. Flavoproteins that are fully reduced (two electrons accepted) generally have an absorption maximum near 360 nm. When partially reduced (one electron), they acquire another absorption maximum at about 450 nm when fully oxidized, the flavin has maxima at 370 and 440 nm. The intermediate radical form, reduced by one electron, has absorption maxima at 380, 480, 580, and 625 nm. These changes can be used to assay reactions involving a flavoprotein. 

The spectra of adrenodoxin and testis iron protein reduced either by dithionite or by NADPH plus adrenodoxin reductase (flavoprotein) exhibit a distinct feature (Fig. 2) the absorption maxima at 455 mp, 414 mp, and 320 mp graetly decrease, although they do not disappear completely. A new distinct maximum appears at 540 mp. However, adrenodoxin can not be reduced by ascorbate or borohydride. The chemical reduction by an excess amount of dithionite gives an 86% decrease in the absorbance, whereas the enzymatic reduction produces a 65% decrease. This difference can be interpreted as gradual loss of iron from the chromophore upon reduction by dithionite. When the reduced form is reoxidized by air, the original spectrum can be regained. The identical characteristic is observed in preparations of testis iron protein. Therefore, adrenal and testis non-heme iron proteins are autoxidizable as well as bacterial and spinach ferredoxins are. 

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