The Ketoreduction and Cyclization of Aromatic Polyketide Biosynthesis

芳香族聚酮生物合成的酮还原和环化

• 批准号: 7827277
• 负责人: Shiou-Chuan Tsai
• 金额: $ 12.63万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2012-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7827277
• 关键词: Active Sites; Affect; Affinity; Amino Acid Sequence; Anabolism; Antibiotics; Aromatase; Bicyclo Compounds; Binding; Biological; Biological Factors; CYP19A1 gene; Catalysis; Chemicals; Cloning; Communities; Complement component C1s; Complex; Crystallization; Cyclization; DNA Sequence; Data; Dehydration; Development; Docking; Doxorubicin; Engineering; Environment; Enzyme Interaction; Enzyme Kinetics; Enzymes; Equipment; Estrogen Antagonists; Family; Fatty Acids; Fatty-acid synthase; Figs - dietary; Funding; Gene Cluster; Genes; Grant; Hydro-Lyases; In Vitro; Individual; Investigation; Kinetics; Knowledge; Label; Laboratories; Length; Libraries; Ligands; Link; Macrolide Antibiotics; Modification; Molecular; Multienzyme Complexes; Mutagenesis; Mutate; Mutation; Occupations; Outcome; Parents; Pattern; Peptide Sequence Determination; Pharmacologic Substance; Plicamycin; Population; Positioning Attribute; Principal Investigator; Protein Engineering; Proteins; Public Health; Recovery; Reporting; Research; Research Personnel; Sequence Homology; Simulate; Specificity; Stepparent; Streptomyces; Structure; Structure-Activity Relationship; Substrate Specificity; System; Testing; Tetracyclines; Text; TimeLine; Training; Translating; United States National Institutes of Health; Variant; X-Ray Crystallography; actinorhodin; antitumor agent; base; combinatorial; crosslink; enzyme activity; genetic analysis; graduate student; in vitro Assay; in vivo; innovation; molecular assembly/self assembly; mutant; novel; polyketide synthase; protein complex; protein protein interaction; public health relevance; skills; small molecule; stereochemistry; sugar; tool; tyrosyl-lysine

DESCRIPTION (provided by applicant):This R01 revision application answers RFA NOT-OD-09-058 (Notice Title: NIH Announces the Availability of Recovery Act Funds for Competitive Revision Applications). This is a revision application for the R01 grant GM076330, titled "The Ketoreduction and Cyclization of Aromatic Polyketide Biosynthesis", whose specific aims were 1. Determine the sequence-structure-function relationship of ketoreductase (KR) that leads to its unique reduction specificity. 2. Determine the sequence-structure-function relationship of aromatase/cyclase (ARO/CYC) that leads to its unique cyclization specificity. 3. Determine the importance of protein-protein interactions on the sequence-structure-function relationship between KR and ARO/CYC. We were pleased to report that our progress towards these three aims has exceeded the proposed timelines. In order to further accelerate the research pace towards understanding polyketide ketoreduction and cyclization, we proposed the following three aims that do not overlap, but rather complementary to the parent R01 aims: AIM 1. Determine the molecular basis of ketoreduction using chemical probes. AIM 2. Determine the molecular basis of aromatic polyketide cyclization using chemical probes. AIM 3. Solve crystal structures of chemically cross-linked multi-domain PKS complexes. The proposed research is significant, because the outcome will answer important questions about how polyketide reduction and cyclization are precisely controlled. It is also innovative by providing new information about KR and ARO/CYC at a molecular level not achieved previously. The long-term biomedical relevance is that the polyketide research community can apply the sequence-structure-function relationships determined from this proposal to diversify the population of "unnatural" natural products via protein engineering, such that a library of novel polyketides with different ketoreduction and cyclization patterns can be produced. The revision will accelerate the tempo of scientific research on polyketide synthase (PKS) and allow for job creation and retention for three graduate students and the hiring of one new postdoctoral researcher, and procuring laboratory equipments to accelerate the tempo of the proposed research. PUBLIC HEALTH RELEVANCE: This is expected to positively affect public health, because it will allow the development of new "unnatural" natural products that can be screened for new pharmaceutical activities. Meanwhile, the fundamental new knowledge obtained in this proposal on correlating PKS sequence-structure with the catalysis, substrate specificity and protein-protein interactions during polyketide ketoreduction and cyclization is expected to have a high impact on the natural product research communities.

说明（由申请人提供）：此 R01 修订申请回答 RFA NOT-OD-09-058（通知标题：NIH 宣布恢复法案资金可用于竞争性修订申请）。这是R01拨款GM076330的修订申请，标题为“芳香族聚酮化合物生物合成的酮还原和环化”，其具体目标是1.确定酮还原酶（KR）的序列-结构-功能关系，从而导致其独特的还原特异性。 2. 确定芳香酶/环化酶（ARO/CYC）的序列-结构-功能关系，从而产生其独特的环化特异性。 3.确定蛋白质-蛋白质相互作用对KR和ARO/CYC之间的序列-结构-功能关系的重要性。我们很高兴地报告，我们在实现这三个目标方面所取得的进展已经超出了拟议的时间表。为了进一步加快了解聚酮化合物酮还原和环化的研究步伐，我们提出了以下三个目标，这些目标并不重叠，而是与母体 R01 目标互补： 目的 1. 使用化学探针确定酮还原的分子基础。 目的 2. 使用化学探针确定芳香族聚酮化合物环化的分子基础。 目标 3. 解析化学交联的多域 PKS 复合物的晶体结构。拟议的研究意义重大，因为其结果将回答有关如何精确控制聚酮化合物还原和环化的重要问题。它还通过在分子水平上提供以前未达到的关于 KR 和 ARO/CYC 的新信息而具有创新性。长期的生物医学相关性是，聚酮化合物研究界可以应用该提案确定的序列-结构-功能关系，通过蛋白质工程使“非天然”天然产物群体多样化，从而形成具有不同酮还原和功能的新型聚酮化合物库。可以产生环化模式。此次修订将加快聚酮合酶（PKS）科学研究的节奏，并为三名研究生创造和保留就业机会，聘用一名新的博士后研究员，并采购实验室设备以加快拟议研究的节奏。 公共健康相关性：预计这将对公共健康产生积极影响，因为它将允许开发新的“非天然”天然产品，并可筛选新的药物活性。同时，该提案中获得的关于将 PKS 序列结构与聚酮化合物酮还原和环化过程中的催化、底物特异性和蛋白质-蛋白质相互作用相关联的基本新知识预计将对天然产物研究界产生重大影响。