New Vistas in Vitamin K Metabolism

描述（由申请方提供）：本研究的长期目标是确定维生素K拮抗剂和维生素K拮抗剂的综合代谢事件网络，以了解受试者对这种必需膳食营养素的反应差异。在新的应用中，我们将建立P450酶，CYP 4F 2和CYP 4F 11作为关键的维生素K羟化酶，启动这种必需的膳食成分的catalysis。此外，我们将 确定新鉴定的人UBDIA 1酶在维生素K1和K2相互转化中的作用。这些努力将建立核心维生素K循环蛋白和辅助酶之间的联系，这些酶调节驱动Gla蛋白生产的氢醌辅因子的可用性。由于在高度脆弱的缺钾人群（例如患有慢性肾病、癌症患者）中补充维生素K的个体间差异很大，因此这些研究具有重要意义。 本研究计划的目标有三个方面，（i）确定人CYP 4F 2和CYP 4F 11在维生素K催化剂启动中的相对作用，（ii）确定UBIAD 1在维生素K1转化为K2（MK-4）中的作用和化学作用机制，和（iii）确定这三种基因中每一种的共同遗传变异对人体组织和日益复杂的细胞模型中维生素K稳态的影响，作为未来体内研究的先驱。 在初步研究中，我们已经完成了三个目标酶的异源表达的重要任务，并开发了高灵敏度和特异性的串联MS分析方法，用于分析各种生物基质中的维生素K。有了这些工具，我们建议在Aim 1中评估CYP 4F 2和CYP 4F 11在维生素K催化剂中的作用。在目标2中，我们将表达和纯化人UBIAD 1，以明确评估酶的催化能力和维生素K1转化为K3的化学机制。在目标3中，我们将使用库存的，基因分型的人肝组织和一种新的工程细胞系统，用于分析Gla蛋白的产生，以确定这些新描述的维生素K周期相关基因中常见的遗传变异对细胞维生素K稳态的影响。

DESCRIPTION (provided by applicant): The long-term aim of this research is to define the network of integrated metabolic events that underlie vitamin K anabolism and catabolism, in order to understand inter-subject variability in response to this essential dietary nutrient. In ths new application, we will establish the P450 enzymes, CYP4F2 and CYP4F11 as the key vitamin K hydroxylases that initiate catabolism of this essential dietary constituent. In addition, we will determine the role of the newly identified human UBDIA1 enzyme in the interconversion of vitamins K1 and K2. These efforts will establish the link between the core vitamin K cycle proteins and ancillary enzymes that modulate the availability of the hydroquinone cofactors that drive Gla protein production. Because there is a high degree of inter-individual variability in response to supplementation with vitamin K in highly vulnerable K-deficient populations (e.g. those suffering from chronic kidney disease, cancer patients), these studies are of high significance. The goals of this research proposal are three-fold, (i) to establish the relative roes of human CYP4F2 and CYP4F11 in the initiation of vitamin K catabolism, (ii) to establish the role and chemical mechanism of action of UBIAD1 in conversion of vitamin K 1 to K2 (MK-4), and (iii) to ascertain the influence of common genetic variation in each of these three genes on vitamin K homeostatis in human tissues and cellular models of increasing complexity, as a precursor to future in vivo studies. In preliminary studies, we have accomplished the important tasks of heterologous expression of each of the three target enzymes, and development of highly sensitive and specific tandem MS assays for analysis of vitamin K in a variety of biological matrices. With these tools in hand, we propose, in Aim1, to evaluate the roles of CYP4F2 and CYP4F11 in vitamin K catabolism. In Aim 2, we will express and purify human UBIAD1 to enable an unambiguous assessment of the enzyme's catalytic capability and chemical mechanism for conversion of vitamin K1 to K3. In Aim 3, we will use banked, genotyped human liver tissue and a novel engineered cell system for analysis of Gla protein production to determine the effect of common genetic variation in these newly described vitamin K cycle-associated genes on cellular vitamin K homeostasis.