Günzler V, Hanauske-Abel H M, Myllylä R, Kaska D D, Hanauske A, Kivirikko K I
Collagen Research Unit, University of Oulu, Finland.
Biochem J. 1988 Apr 15;251(2):365-72. doi: 10.1042/bj2510365.
The anthracyclines doxorubicin and daunorubicin were found to act as irreversible inhibitors of prolyl 4-hydroxylase. The reaction rate for enzyme from both chick and human origin was first order, the concentration of inhibitor giving 50% inhibition being 60 microM for both compounds after 1 h. The effect was dependent on the presence of iron ions in the reaction mixture. Inactivation could be prevented by addition of high concentrations of ascorbate, but not 2-oxoglutarate, before the inactivation period. The same results were obtained with competitive analogues of these cosubstrates. Lysyl hydroxylase from chick embryos was also susceptible to inactivation. Its activity was decreased by 50% after incubation for 1 h with a 150 microM concentration of the inhibitors. When chick-embryo prolyl 4-hydroxylase was incubated with [14-14C]doxorubicin, both enzyme subunits were radioactively labelled, about 70% of the total radioactivity being found in the alpha-subunit. Since the anthracyclines are known to undergo a redox reaction generating semiquinone radicals with Fe3+ only, the results suggest that the enzyme-bound iron ion is oxidized to a tervalent intermediate in uncoupled reaction cycles. The data also suggest that both enzyme subunits contribute to the catalytic site of prolyl 4-hydroxylase.
发现蒽环类药物阿霉素和柔红霉素可作为脯氨酰4-羟化酶的不可逆抑制剂。来自鸡和人类的该酶的反应速率均为一级反应，1小时后两种化合物产生50%抑制作用的抑制剂浓度均为60微摩尔。该作用取决于反应混合物中铁离子的存在。在失活期之前加入高浓度的抗坏血酸可防止失活，但加入2-氧代戊二酸则不能。这些共底物的竞争性类似物也得到了相同的结果。鸡胚赖氨酰羟化酶也易被失活。用150微摩尔浓度的抑制剂孵育1小时后，其活性降低了50%。当用[14-14C]阿霉素孵育鸡胚脯氨酰4-羟化酶时，两个酶亚基均被放射性标记，约70%的总放射性存在于α亚基中。由于已知蒽环类药物仅与Fe3+发生氧化还原反应生成半醌自由基，结果表明在未偶联的反应循环中，酶结合的铁离子被氧化为三价中间体。数据还表明两个酶亚基均对脯氨酰4-羟化酶的催化位点有贡献。
