Characterization of Vitamin K Epoxide Reductase

维生素 K 环氧化物还原酶的表征

• 批准号: 7179243
• 负责人: DARREL W STAFFORD
• 金额: $ 35.45万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-03-01 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7179243
• 关键词: Amino Acids; Antibodies; Binding Sites; Blood coagulation; Cells; Coagulants; Cysteine; Disease; Dose; Fright; Funding; Genes; Genetic Polymorphism; Grant; Hemorrhage; Laboratories; Lead; Mass Spectrum Analysis; Physiological; Post-Translational Protein Processing; Production; Property; Protein Isoforms; Recombinants; Research; Research Personnel; Role; Site; Stroke; System; Vitamin K; Vitamin K Reductase; Warfarin; bone metabolism; carboxylation; design; glycosylation; prevent; response; vitamin K epoxide reductase

DESCRIPTION (provided by applicant): The long-term objective of this project is to understand the enzymatic properties of the vitamin K epoxide reductase (VKOR) and its role in vitamin K-dependent carboxylation. Carboxylation is an essential post-translational modification for proteins important in several physiologic functions, including blood coagulation and bone metabolism. Recently, researchers in this laboratory identified the warfarin-sensitive VKOR gene. VKOR is the target of warfarin, the most widely prescribed anti-coagulant for thromboembolic disorders. Although estimated to prevent twenty strokes per induced bleeding episode, warfarin is under-utilized because of fear of bleeding. Research funded by this grant hopefully will lead to therapies that will alleviate some of the problems involved in warfarin treatment. Specifically, we propose to accomplish the following during the tenure of this grant: (1) express the isoforms of the VKOR gene to determine which have activity; (2) purify the VKOR using the amino acid tag we have inserted and the antibody to this tag; (3) investigate different cell expression systems to determine which is best for production of recombinant VKOR; (4) investigate the status of cysteines in VKOR, the glycosylation sites and other potential post-translational modifications, and the warfarin binding site by mass spectrometry; and (5) investigate the effect of vitamin K epoxide reductase (VKOR) gene polymorphisms on response to warfarin. This study of VKOR should lead to a better understanding of the mechanism of warfarin inhibition. It may also result in more accurate dosing of warfarin and in the design of new anti-coagulants.

描述（由申请人提供）：该项目的长期目的是了解维生素K环氧化氧化物还原酶（VKOR）及其在维生素K依赖性羧化中的作用。羧化是在几种生理功能（包括血液凝结和骨代谢）中重要的蛋白质的基本翻译后修饰。最近，该实验室的研究人员确定了对华法林敏感的VKOR基因。 VKOR是华法林（Warfarin）的目标，华法林（Warfarin）是血栓栓塞性疾病最广泛规定的抗凝蛋白剂。尽管据估计，由于担心出血，华法林估计每次引起的出血性发作二十个中风，但未充分利用。这项赠款资助的研究有望导致疗法，以减轻华法林治疗中涉及的一些问题。具体而言，我们建议在本赠款任期期间完成以下内容：（1）表达VKOR基因的同工型以确定哪些活性； （2）使用已插入的氨基酸标签和该标签的抗体纯化VKOR； （3）研究不同的细胞表达系统，以确定哪种最适合重组VKOR的生产； （4）研究半胱氨酸在VKOR中的状态，糖基化位点和其他潜在的翻译后修饰以及通过质谱法进行华法林结合位点； （5）研究维生素K环氧化物还原酶（VKOR）基因多态性对华法林反应的影响。对VKOR的这项研究应可以更好地理解华法林抑制的机制。这也可能导致华法林的更准确剂量和新的抗凝剂的设计。