One-Step Synthesis of D-Amino Acids

D-氨基酸的一步合成

• 批准号: 6832095
• 负责人: SCOTT J NOVICK
• 金额: $ 10万
• 依托单位: BIOCATALYTICS, INC.
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-01 至 2005-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6832095
• 关键词: aminoacid; biotechnology; chemical kinetics; chemical synthesis; cofactor; directed evolution; enzyme substrate; genetic library; genetic screening; mutant; oxidoreductase; polymerase chain reaction; stereoisomer; technology /technique development

DESCRIPTION (provided by applicant): Many currently produced drugs such as beta-Iactam antibiotics, thrombin inhibitors and fertility treatments, and fine chemicals such as insecticides contain D-amino acids. Some of these compounds are produced on a scale greater than 5000 tons/yr. In addition, there are a number of D-amino acid containing drugs, either in the research or clinical test stages, used for the treatment of such illnesses as endometriosis and uterine fibroids in women, benign prostatic hypertrophy in men, and HIV. D-Amino acids have the advantage of often being more bioactive than their L-counterpart and they are also frequently more stable as they are less likely to undergo enzymatic degradation in the liver, kidney, and bloodstream. Clearly, there is significant demand for enantiomerically pure D-amino acids and this is expected to rise significantly in the near future as they become critical components of new pharmaceuticals. While current technologies exist for producing D-amino acids they all have drawbacks including poor yields and the requirement of multiple reaction steps. New methods to improve upon current technologies are needed. One method, as discussed in this proposal, is to produce D-amino acids from the 2-keto acid precursor using a D-amino acid dehydrogenase. This will allow for the single-step synthesis of D-amino acid from inexpensive starting materials. To this end, it is proposed to use direct evolution to generate broadrange D-amino acid dehydrogenases. The starting enzyme, meso-2,6-diaminopimelate-D-dehydrogenase (DAPDH) is highly selective for the synthesis of D-amino acids. The specific aims of this proposal are to generate a library of mutant DAPDH, screen this library for activity towards important D-amino acids, characterize the best mutants, and use these mutants to perform a gram scale synthesis of one or more key D-amino acids.

描述（由申请人提供）：目前生产的许多药物（如β-内酰胺类抗生素、凝血酶抑制剂和生育治疗药物）和精细化学品（如杀虫剂）均含有D-氨基酸。这些化合物中的一些以大于5000吨/年的规模生产。此外，有许多含D-氨基酸的药物，无论是在研究或临床试验阶段，用于治疗妇女的子宫内膜异位症和子宫肌瘤，男性的良性前列腺肥大和艾滋病毒等疾病。D-氨基酸的优点是通常比它们的L-对应物更具生物活性，并且它们通常也更稳定，因为它们不太可能在肝脏，肾脏和血液中经历酶降解。显然，对映体纯D-氨基酸的需求量很大，预计在不久的将来，随着它们成为新药的关键组分，需求量将显著上升。 虽然目前存在用于生产D-氨基酸的技术，但它们都具有缺点，包括产率低和需要多个反应步骤。需要新的方法来改进现有的技术。如本提案中所讨论的，一种方法是使用D-氨基酸脱氢酶从2-酮酸前体生产D-氨基酸。这将允许从廉价的起始材料一步合成D-氨基酸。为此，建议使用直接进化来产生宽范围的D-氨基酸内切酶。起始酶内消旋-2，6-二氨基庚二酸-D-脱氢酶（DAPDH）对D-氨基酸的合成具有高度选择性。该提议的具体目的是产生突变DAPDH文库，筛选该文库对重要D-氨基酸的活性，表征最佳突变体，并使用这些突变体进行一种或多种关键D-氨基酸的克级合成。