Activation of androgen biosynthesis and drug metabolism by cytochrome b5

所有的19碳雄激素都是由21碳类固醇通过连续的17？羟基酶产生的。 细胞色素P450c17(CYP17A1)的17，20-裂解酶活性。复杂的化学 细胞色素b5(B5)选择性地刺激17，20-裂解酶反应10倍。 在试管中。与b5与其他一些细胞色素P450的相互作用不同，P450 显然涉及b5的电子转移，我们的数据认为b5是变构的 激活细胞色素P17A1的17，20-裂解酶活性。我们已经确定了b5的一个特定区域。 这对于刺激17，20-裂解酶的活性是至关重要的，并且已经表明 这种刺激是底物依赖的。我们现在建议澄清一下 B5在体内对17，20-裂解酶活性和雄激素合成的作用 比较b5-CyP17A1与b5相互作用的机制和结构特征 对其他P450介导的反应的作用，为详细询问 B5对细胞色素P17A1的作用机制。 为了首先证明b5对雄激素的生物合成是必不可少的，我们将产生和 为目标1描述睾丸中缺乏b5的小鼠。我们将描述其生育力， 荷尔蒙的产生和这些小鼠的睾丸组织学。该酶的酶活性 我们将对这些睾丸进行详细研究。在目标2中，我们将确定b5上的残基是否 刺激17，20-裂解酶活性所需的物质也是调节 并开始测定其他细胞色素P450的活性是否相同 所有的P450酶都涉及到微观步骤。因此，我们将定义 B5对细胞色素P17A1的作用在哺乳动物生理学中的重要性，我们将开始 确定其作用机制是否类似于b5对其他细胞色素P450的作用。 这一知识将为抑制雄激素的产生提供一种新的方法，通过 靶向CYP17a1-b5相互作用。

All 19-carbon androgens derive from 21-carbon steroids via sequential 17¿-hydroxylase and 17,20-lyase activities of cytochrome P450c17 (CYP17A1). The complex chemistry of the 17,20-lyase reaction is selectively stimulated up to 10-fold by cytochrome b5 (b5) in vitro. In contrast to the interactions of b5 with some other cytochromes P450, which apparently involve electron transfer from b5, our data argue that b5 allosterically activates the 17,20-lyase activity of CYP17A1. We have identified a specific region of b5 that is critical for stimulation of 17,20-lyase activity and have shown that the magnitude of this stimulation is substrate-dependent. We now propose to elucidate the importance of this action of b5 on 17,20-lyase activity and androgen synthesis in vivo and to compare the mechanistic and structural features of the b5-CYP17A1 interaction with b5 action on other P450-mediated reactions, in preparation for a detailed interrogation of the mechanism of b5 action on CYP17A1. To first prove that b5 is essential for androgen biosynthesis, we will generate and characterize mice lacking b5 in the testis for Aim 1. We will characterize the fertility, hormone production, and testicular histology of these mice. The enzymatic activity of the testes will be studied in detail. In Aim 2, we will determine if residues on b5 that are required for stimulating 17,20-lyase activity are also essential for modulating the activities of other cytochromes P450 and begin to determine whether the same microscopic steps are involved for all P450 enzymes. We thus will define the importance of b5 action on CYP17A1 in mammalian physiology, and we will begin to ascertain if the mechanism of action is similar to b5 action on other cytochromes P450. This knowledge will provide a novel approach for suppressing androgen production, by targeting the CYP17A1-b5 interaction.