In vitro glycorandomization of natural products

描述（由申请人提供）：糖附着在天然存在和/或合成的小分子上可以显著影响相应的机制、药效学、药代动力学甚至亲本结构的专利寿命。然而，由于与传统糖基化策略相关的技术挑战，糖基化在药物发现和/或开发背景下的应用仍未得到充分探索。该项目的首要目标是开发简单且用户友好的化学酶方法，用于靶支架的差异糖基化，最终实现此类药物的发现/开发探索。为了实现这一目标，本研究的第一阶段（1-5年）对化学酶糖随机化（一锅三酶策略，能够激活和连接游离单糖到复杂的天然产物支架上）的概念进行了关键的验证，并提供了关于两个关键但知之甚少的酶类（糖聚糖激酶和糖-1-磷酸核苷酸转移酶）的基本信息。该项目的第二阶段（6-10年级）极大地扩展了第一代系统对各种支架的尝试应用，在许多情况下取得了成功，但也暴露了平台的关键局限性。重要的是，在第二阶段进行的工作也导致了关于糖基转移酶（GT）催化反应的基本新知识，作为启动本研究第三阶段的基础。具体来说，本研究的第三阶段（11-15年级）需要

DESCRIPTION (provided by applicant): The attachment of sugars to naturally-occurring and/or synthetic small molecules can dramatically influence the corresponding mechanism, pharmacodynamics, pharmacokinetics and even patent life of the parental structure. Yet, due to the technical challenges associated with conventional glycosylation strategies, the application of glycosylation in the context of drug discovery and/or development remains underexplored. An overarching aim of this program has been to develop simple and user-friendly chemoenzymatic methods for the differential glycosylation of target scaffolds to ultimately enable such drug discovery/development exploration. Toward this goal, the first phase of this study (years 1-5) led to the key proof of concept for chemoenzymatic glycorandomization (a one pot, three enzyme strategy capable of activating and attaching free monosaccharides to complex natural product scaffolds) and also provided fundamental information regarding two critical, but poorly understood, enzyme classes (anomeric sugar kinases and sugar-1-phosphate nucleotidylyltransferases). The second phase of this project (years 6-10) dramatically expanded the attempted application of this first generation system toward a range of diverse scaffolds, leading to success in many cases but also exposing key limitations of the platform. Importantly, the work conducted during the second phase also led to fundamental new knowledge regarding glycosyltransferase (GT)-catalyzed reactions that serves as the basis from which to launch the third phase of this study. Specifically, the proposed third phase of this study (years 11-15) takes advantage of our recent abilities to evolve highly permissive GTs and also drive GT-catalyzed reactions in reverse to enable next generation single enzyme or dual enzyme transglycosylation strategies for differential glycosylation of an array of structurally-diverse scaffolds (including natural product-based or synthetic, glycosylated or non-glycosylated, parental scaffolds). We propose to take full advantage of the current state of the art to: i) narrow the gaps of knowledge in understanding functional GT structure-activity-relationships; ii) specifically develop a range o catalysts for the production of novel sugar nucleotides (anticipated to be of broad use to the glycobiology community); iii) develop a range of catalysts for the differential glycosylation of a key set of structurally diverse anti-infective and anticancer scaffolds; and iv)exploit the corresponding differentially glycosylated scaffolds as a novel source for the discovery of new antiinfective and anticancer leads. PUBLIC HEALTH RELEVANCE: We propose to develop simple and user-friendly chemoenzymatic methods for the differential glycosylation of complex target scaffolds to enable glycoconjugation as a tool for drug discovery and development. The models selected as part of the current study are anticipated to hold particularly high potential for the discovery of new antibacterial, antiviral and anticancer leads.