REGULATION OF LIVER CYTOCHROME P450 TURNOVER/HEPATIC DEGRADATION OF P450 ENZYMES

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心赠款提供。次级项目的主要支助 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 表示子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 肝微粒体血红素蛋白细胞色素P450（P450）包括多种组成型和诱导型酶。它们在各种生理学相关的内源性和外源性物质的氧化/还原代谢中起作用。这些底物可以通过转录/翻译机制诱导各种P450的含量，但也可以通过蛋白质稳定，即抑制其蛋白水解降解。 这种降解通过泛素依赖性蛋白酶体降解（UPD）、自噬-溶酶体降解（ALD）或在某些情况下通过两种途径进行。 我们研究的长期目标集中在分子水平上表征这些过程：标记这些蛋白质用于细胞处置的降解决定子和翻译后修饰以及其中涉及的分子伴侣。具体而言，使用质谱和/或蛋白质组学分析，我们希望表征P450泛素化的位点，分子伴侣和参与其降解的因素。研究正在与Burlingame，Medzihradszky和Maltby博士合作，以确定P450蛋白的泛素化的精确氨基酸残基。同时，我们的研究也集中在血红素在调节肝脏P450合成和周转的作用。拟议的研究集中在P450生物学的生理相关但被忽视的方面。 由于P450是整合ER膜蛋白，阐明其营业额将提供其他ER居民的生物原型。此外，这些研究集中在P450 3A 4，主要的人类肝脏和肠道酶及其大鼠直系同源物，负责超过60%的临床处方药物的代谢，并且特别容易受到这种生物学命运的影响。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The hepatic microsomal hemoproteins cytochromes P450 (P450) include multiple constitutive and inducible enzymes. that are instrumental in the oxidative/reductive metabolism of various physiologically relevant endobiotics and xenobiotics. These substrates can induce the content of various P450s via transcriptional/translational mechanisms but also via protein stabilization i.e. inhibition of its proteolytic degradation. This degradation proceeds via ubiquitin-dependent proteasomal degradation (UPD), autophagic-lysosomal degradation (ALD) or in some cases by both pathways. The long-term goals of our research are centered on the characterization of these processes at the molecular level: the degrons and posttranslational modifications that mark these proteins for cellular disposal and the molecular partners involved therein. Specifically, using mass spectrometric and/or proteomic analyses we wish to characterize the sites of P450 ubiquitination, the chaperones and factors involved in their degradation. Studies are in progress in collaboration with Drs. Burlingame, Medzihradszky and Maltby to identify the precise amino acid residue of the P450 protein that is ubiquitinated. In parallel our studies also focus on the role of heme in regulating the synthesis and turnover of hepatic P450s. The proposed studies center on a physiologically relevant but neglected aspect of P450 biology. Because P450s are integral ER-membrane proteins, elucidation of its turnover will provide a biological prototype for other ER-residents. Furthermore, these studies are focussed on P450 3A4, the major human liver and intestinal enzyme and its rat orthologs, which are responsible for the metabolism of over 60% of clinically prescribed drugs, and are particularly susceptible to this biological fate.