The Biochemistry of CAD

CAD 的生物化学

• 批准号: 9105826
• 负责人: Jeffrey Davidson
• 金额: $ 25.4万
• 依托单位: University of Kentucky Research Foundation
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-08-15 至 1995-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9105826&HistoricalAwards=false
• 关键词: Biochemistry; CAD

The first three enzymatic steps in the de novo synthesis of pyrimidines (carbamyl phosphate synthetase II, aspartate transcarbamylase, dihydroorotase) are the same in all cells, whether prokaryotic or eukaryotic. The structure, organization, and regulation of these three enzymatic activities and their respective genes vary among species. In bacteria, each enzyme activity is associated with a separate protein. In mammals, they are found together in a single translation product of 240,000 daltons called CAD. By placing portions of hamster CAD cDNA into expression vectors, one can attempt to complement E. coli and hamster cell mutants defective in each of the three enzyme activities. This avenue of experimentation allows one (i) identify the minimum genetic information encoding each domain, (ii) to learn pyrimidine metabolism is affected when specific mutations are introduced that eliminate allosteric regulation of CAD or alter activity of one of its enzymes and (iii) to gain insight into the roles of the bridging sequences that link the enzymatic domains. Combining the approaches of molecular biology, genetics, and biochemistry should provide insight about the structure, function and evolution of a multifunctional protein that plays a key role in an important biosynthetic pathway.

在所有细胞中，无论是原核生物还是真核。 这三种酶活性及其各自基因的结构，组织和调节在物种之间有所不同。 在细菌中，每种酶活性与单独的蛋白质有关。 在哺乳动物中，它们以240,000个Daltons的单一翻译产品一起发现，称为CAD。通过将仓鼠CAD cDNA的一部分放入表达载体中，可以尝试在三种酶活性中的每一个中补充大肠杆菌和仓鼠细胞突变体有缺陷。 这种实验途径允许一（i）确定编码每个领域的最低遗传信息，（ii）在引入特定的突变时，会影响吡霉素代谢，从而消除对CAD的变形或改变其一种酶的活性变为活性的特定突变，并（iii）洞察纤维化序列的角色，以链接Enzymatics的纤维序列。 结合了分子生物学，遗传学和生物化学的方法，应提供有关多功能蛋白的结构，功能和演变的见解，该蛋白在重要的生物合成途径中起着关键作用。