Department of Pharmacology and Experimental Therapeutics and The Stanley S. Scott Cancer Center, Louisiana State University Health Science Center, New Orleans, Louisiana
Department of Pharmacology and Experimental Therapeutics and The Stanley S. Scott Cancer Center, Louisiana State University Health Science Center, New Orleans, Louisiana.
Drug Metab Dispos. 2023 Sep;51(9):1196-1206. doi: 10.1124/dmd.123.001287. Epub 2023 Jun 22.
Liver cytochrome P450s (CYPs) of the endoplasmic reticulum (ER) are involved in the metabolism of exogenous and endogenous chemicals. The ER is not uniform, but possesses ordered lipid microdomains containing higher levels of saturated fatty acids, sphingomyelin, and cholesterol and disordered regions containing higher levels of polyunsaturated fatty acid chains. The various forms of drug-metabolizing P450s partition to either the ordered or disordered lipid microdomains with different degrees of specificity. P450s readily form complexes with ER-resident proteins, including other forms of P450. This study aims to ascertain whether lipid microdomain localization influences protein-P450 interactions in rat liver microsomes. Thus, liver microsomes were co-immunoprecipitated with CYP1A2-specific and CYP3A-specific antibodies, and the co-immunoprecipitating proteins were identified by liquid chromatography mass spectrometry proteomic analysis. These two P450s preferentially partition to ordered and disordered microdomains, respectively. More than 100 proteins were co-immunoprecipitated with each P450. Segregation of proteins into different microdomains did not preclude their interaction. However, CYP3A interacted broadly with proteins from ordered microdomains, whereas CYP1A2 reacted with a limited subset of these proteins. This is consistent with the concept of lipid raft heterogeneity and may indicate that CYP1A2 is targeted to a specific type of lipid raft. Although many of the interacting proteins for both P450s were other-drug metabolizing enzymes, other interactions were also evident. The consistent CYP3A binding partners were predominantly involved in phase I/II drug metabolism; however, CYP1A2 interacted not only with xenobiotic metabolizing enzymes, but also with enzymes involved in diverse cellular responses such as ER stress and protein folding. SIGNIFICANCE STATEMENT: This work describes the protein interactomes in rat liver microsomes of two important cytochromes P450s (CYP1A2 and CYP3A) in drug metabolism and describes the relationship of the interacting proteins to lipid microdomain distribution.
内质网（ER）中的肝细胞色素 P450（CYP）参与外源性和内源性化学物质的代谢。ER 不是均匀的，而是具有有序的脂质微区，其中含有较高水平的饱和脂肪酸、神经鞘磷脂和胆固醇，以及无序区域，其中含有较高水平的多不饱和脂肪酸链。各种形式的药物代谢 CYP 以不同程度的特异性分配到有序或无序的脂质微区。P450 很容易与 ER 驻留蛋白形成复合物，包括其他形式的 P450。本研究旨在确定脂质微区定位是否影响大鼠肝微粒体中蛋白质-P450 相互作用。因此，用 CYP1A2 特异性和 CYP3A 特异性抗体共免疫沉淀肝微粒体，并通过液相色谱-质谱蛋白质组学分析鉴定共免疫沉淀的蛋白质。这两种 P450 分别优先分配到有序和无序微区。用每种 P450 共免疫沉淀了 100 多种蛋白质。蛋白质的分离到不同的微区并不排除它们的相互作用。然而，CYP3A 与来自有序微区的蛋白质广泛相互作用，而 CYP1A2 则与这些蛋白质的有限子集反应。这与脂筏异质性的概念一致，可能表明 CYP1A2 被靶向到特定类型的脂筏。尽管两种 P450 的许多相互作用蛋白都是其他药物代谢酶，但也存在其他相互作用。一致的 CYP3A 结合伴侣主要参与 I/II 期药物代谢；然而，CYP1A2 不仅与外源性代谢酶相互作用，还与参与细胞应激和蛋白质折叠等多种细胞反应的酶相互作用。意义陈述：本工作描述了药物代谢中两种重要细胞色素 P450（CYP1A2 和 CYP3A）在大鼠肝微粒体中的蛋白质相互作用组，并描述了相互作用蛋白与脂质微区分布的关系。
