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In an attempt to elucidate the mechanism(s) behind the susceptibility of tissues to the toxic effects of chemicals, NADPH-cytochrome P450 reductase IgG has been used to map the distribution and localization of the cytochrome P450 system in hepatic and extrahepatk human tissues. Employing the Western blot procedure this antibody recognized a single band in human liver mkrosomes which corresponded in mol. wt to the purified reductase. Immunoreactive NADPH-cytochrome P450 reductase staining was detected in all zones of the liver acinus, with maximal staining in hepatocytes adjacent to the terminal hepatic venules (zone 3). Considerable variation in the intensity of reductase staining was observed in different segments of the gastrointestinal tract. Staining was most intense in the enterocytes of the small intestine, with maximal staining at the tips of the villi. Colonic epithelial cells were variably positive while the rectum was negative. Pancreatic ductal cells were positive whereas exocrine cells were negative. In the lung, reductase was detected in bronchiolar and bronchial epithelial cells, Clara cells and alveolar lining cells. In the kidney, the proximal and distal convoluted tubules, the loops of Henle, the collecting ducts in the medulla and the transitional epithelium all stained positively for reductase. The results demonstrate specific cellular localization of NADPH-cytochrome P450 reductase, and hence the cytochrome P450 system, in human tissues. The differential distribution of the reductase within human tissues may lead to a better understanding of the mechanism underlining site-specific carcinogenesis. [ABSTRACT FROM PUBLISHER]

1. The effects of temperature and ligands (AMP, IMP and glucose 1-phosphate) on the properties of rabbit muscle phosphorylase b have been examined by several methods. 2 The non-covalently bound fluorescent probe 2-methylanilino naphthalene 6-sulphonate is shown to detect the dimer to tetramer transition induced by AMP The aggregation is slow and has a second order rate constant of 3.1 × 105 M-1 min-1. 3. The interaction of Mn2+ with phosphorylase b is studied by proton relaxation enhancement and electron spin resonance, The limitations of graphical procedures for analysing proton relaxation enhancement data are discussed and better iterative procedures are presented. 4. There are two equivalent and independent binding sites for Mn2+ per phosphorylase b dimer. These sites (Kd = 0.17 mM) have a 30-fold higher affinity for Mn2+ than those involved in tetramer formation as measured by methylanilino naphthalene sulphonate fluorescence. 5. The reaction of a specific sulphydryl reagent, 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole with phosphorylase b can be separated into sets of one fast and two slowly reacting groups per protomer. Modification of one of the slow groups results in inactivation. 6. The temperature dependences of the proton relaxation rates and of the pattern of - SH reactivity discontinuities at around 13 °C, This has been interpreted as a change in the enzyme conformation. 7. AMP increases the limiting proton relaxation enhancement from 10.9 to 14 and decreases the reactivity with 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole of the slow - SH groups. The increase in the limiting proton relaxation enhancement is not due to aggregation but is likely to be a result of a ligand induced conformational change. 8. Glucose 1-phosphate decreases the limiting proton relaxation enhancement from 10.9 to 5.8 and increases the reactivity of one of the slow thiols. [ABSTRACT FROM AUTHOR]