Dean P A, Rettie A E, Turnblom S M, Namkung M J, Juchau M R
Chem Biol Interact. 1986 Apr;58(1):79-94. doi: 10.1016/s0009-2797(86)80088-6.
Additions of micromolar concentrations of hematin to washed rat pulmonary microsomal preparations resulted in marked (5-7-fold) increases in the NADPH-dependent generation of phenolic metabolites of benzo[a]pyrene (BaP). 9-Hydroxy-BaP was identified as the major reaction product. Additions of pulmonary cytosolic fractions to microsomes produced no measurable effect but cytosol and hematin added together elicited 25-30-fold increases in total phenolic products. Cytosolic fractions from other tissues, including rat kidneys and perfused rat livers, were also highly effective in enhancing the hematin-mediated increases in monooxygenase activity. However, cytosol from human placental tissues was only minimally effective when either pulmonary or placental microsomes were utilized as enzyme source. Superoxide dismutase and catalase (alone or in combination) had no measurable effect on hematin-mediated increases. Horseradish peroxidase effectively inhibited the hematin-dependent reactions but hematin-independent reactions were inhibited with equal effectiveness. Carbon monoxide profoundly inhibited all hematin-mediated increases in metabolite formation. The activating cytosolic component was non-dialyzable, inactivated by trypsin and heat, and eluted in the void volume from Sephadex G-150 columns. This suggested that the cytosolic factor(s) responsible for the increased hematin-dependent oxidation was a protein(s) with a high molecular weight or perhaps an aggregate or oligomer of proteinaceous material. HPLC profiles indicated a major effect on the generation of phenolics; quinones were also increased but only minimal increases in diols were observed. Results were consistent with the hypothesis that hematin-mediated increases in pulmonary monooxygenase activity result from an increased association of a small pool of pulmonary P-450-apoprotein(s) with the hematin prosthetic group to result in increased levels of an unidentified holocytochrome(s) with a relatively high substrate turnover number. The current data suggest a quaternary interaction among P-450 apoprotein(s), heme prosthetic group, reaction products (particularly 3-hydroxy-BaP) and a cytosolic protein(s). We postulate that the mechanism of action of the cytosolic factor is to facilitate the interaction of hematin with the apocytochrome.
向洗涤过的大鼠肺微粒体制剂中添加微摩尔浓度的血晶素，导致苯并[a]芘（BaP）的NADPH依赖性酚类代谢产物生成显著增加（5 - 7倍）。9 - 羟基 - BaP被鉴定为主要反应产物。向微粒体中添加肺胞质部分未产生可测量的影响，但胞质溶胶和血晶素一起添加会使总酚类产物增加25 - 30倍。来自其他组织的胞质部分，包括大鼠肾脏和灌注的大鼠肝脏，在增强血晶素介导的单加氧酶活性增加方面也非常有效。然而，当以肺或胎盘微粒体作为酶源时，人胎盘组织的胞质溶胶效果甚微。超氧化物歧化酶和过氧化氢酶（单独或联合使用）对血晶素介导的增加没有可测量的影响。辣根过氧化物酶有效地抑制了血晶素依赖性反应，但对非血晶素依赖性反应的抑制效果相同。一氧化碳强烈抑制了所有血晶素介导的代谢产物形成增加。激活的胞质成分不可透析，被胰蛋白酶和加热灭活，并从Sephadex G - 150柱的空体积中洗脱。这表明负责增加血晶素依赖性氧化的胞质因子是一种高分子量的蛋白质，或者可能是蛋白质物质的聚集体或寡聚体。HPLC图谱表明对酚类物质的生成有主要影响；醌类也增加了，但二醇类仅观察到最小程度的增加。结果与以下假设一致，即血晶素介导的肺单加氧酶活性增加是由于一小部分肺P - 450脱辅基蛋白与血晶素辅基的结合增加，导致具有相对高底物周转率的未鉴定全细胞色素水平升高。目前的数据表明P - 450脱辅基蛋白、血红素辅基、反应产物（特别是3 - 羟基 - BaP）和一种胞质蛋白之间存在四级相互作用。我们推测胞质因子的作用机制是促进血晶素与脱细胞色素的相互作用。
