S. coelicolor P450s: Structure/Function/Engineering

S. coelicolor P450s：结构/功能/工程

• 批准号: 6827209
• 负责人: MICHAEL R WATERMAN
• 金额: $ 40.21万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-01 至 2008-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6827209
• 关键词: Streptomyces; X ray crystallography; bacterial genetics; crystallization; cytochrome P450; enzyme structure; enzyme substrate; gene expression; genome; liquid chromatography mass spectrometry; oxygenases; polymerase chain reaction

DESCRIPTION (provided by applicant): Streptomycetes are bacteria known to inhabit the soil and marine ecosystems worldwide. Within their niches streptomycetes have developed complex and efficient mechanisms of defense against other organisms through the production of anti-bacterial, anti-fungal and anti-nematode compounds and other bioactive compounds, many of which have value in human and veterinary medicine. Streptomycetes also exhibit a broad-based ability to detoxify organic poisons. Both these defense systems involve monooxygenases of the cytochrome P450 (CYP) superfamily representing about 0.25% of streptomycete genes. Many of the CYP gene families overlap between Streptomyces spp. as judged by genomic data, but different subfamilies and families are apparent that reflect the different needs in secondary metabolism. The diversity of P450s reflects the selective pressures allowing subtly different activities to evolve and be retained and a long term goal of this project is to understand general features of substrate-binding to streptomycete P450s and strategies used in different species to redesign P450 primary sequence leading to different natural products. This information will establish a knowledge base from which we can consider the production of new and potent bioactive compounds. The genome of Streptomyces coelicolor A3 (2) contains 18 CYP genes and this current application proposes to establish the function of eight of these CYPs selected from our lead-up work, to correlate their high resolution X-ray structure with function, and to generate modified forms of these CYPs by mutagenesis to examine altered biological activity. S. coelicolor is a particularly good system for this study because a large number of its secondary metabolites are known and they represent a diverse group of organic molecules, but also due to the facile molecular systems established for study of this species. Further, most of the CYPs fall into small groups of related enzymes (same family or subfamily). Three laboratories with overlapping expertise and ongoing collaborations have joined forces to achieve these goals. When complete, this study of CYPs from S. coelicolor will set the stage for alteration of other Streptomyces spp. which through modified defense systems can produce new antibiotics and other drugs for human and animal medicine.

描述（由申请人提供）：链霉菌是已知的细菌，可以居住在全球的土壤和海洋生态系统中。在他们的壁ni中，链霉菌通过生产抗细菌，抗真菌和抗新虫化合物以及其他生物活性化合物，开发了针对其他生物体的复杂有效的防御机制，其中许多化合物在人类和兽医医学中具有价值。链霉菌还表现出广泛的有机毒物排毒能力。这两种防御系统都涉及细胞色素P450（CYP）的单加氧酶，代表约0.25％的链霉菌基因。许多CYP基因家族在链霉菌属之间重叠。根据基因组数据的判断，但是不同的亚家族和家庭显然反映了次级代谢的不同需求。 P450的多样性反映了选择性压力，允许不同的活动发展和保留，该项目的长期目标是了解链霉菌链霉菌P450的底物结合的一般特征，以及在不同物种中使用的策略来重新设计P450 P450主要序列，从而导致不同的天然产物。这些信息将建立一个知识库，我们可以从中考虑产生新的和有效的生物活性化合物。 Coelicolor A3（2）的基因组包含18个CYP基因，目前的应用建议建立从我们的铅工作中选择的八个CYP的功能，以将其高分辨率X射线结构与功能相关联，并通过Mutagenesens通过Mutagenesis来检查改变的生物学活性。 Coelicolor S. coelicolor是该研究的特别好的系统，因为它的大量二级代谢产物是已知的，它们代表了多样化的有机分子，但也是由于建立了用于研究该物种的便捷分子系统。此外，大多数CYP分为一小部分相关酶（同一家族或亚家族）。拥有重叠的专业知识和持续合作的三个实验室已经结合起来，以实现这些目标。完成后，对Coelicolor链球菌CYP的这项研究将为改变其他链霉菌属的阶段奠定基础。通过改良的防御系统可以生产新的抗生素和其他药物用于人类和动物医学​​。