Reagents for Chemical Oligophosphorylation, Synthesis of Oligophosphate-Organic Molecule Conjugates, and Biochemical Studies

用于化学低磷酸化、低磷酸盐-有机分子缀合物的合成以及生化研究的试剂

• 批准号: 9886568
• 负责人: CHRISTOPHER C CUMMINS
• 金额: $ 29.17万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9886568
• 关键词: Active Sites; Aldehydes; Amino Acids; Binding; Biochemical; Biological; Cations; Cattle; Cell physiology; Chemicals; Chemistry; Collaborations; Coupling; Crystallization; Crystallography; Development; Diagnostic; Family; Health; High Pressure Liquid Chromatography; Human; Hydroxides; Length; Letters; Lipids; Maps; Medicine; Methodology; Modernization; Nucleosides; Pancreatic ribonuclease; Pathway interactions; Pentas; Peptides; Periodicity; Pharmacologic Substance; Phosphorylation; Physiological; Physiology; Play; Polyps; Post-Translational Protein Processing; Preparation; Proteins; Publications; RNA; Rain; Reaction; Reagent; Reporting; Research Personnel; Ribonucleases; Role; Route; Structure; base; biochemical tools; crystallinity; design; improved; inhibitor/antagonist; inorganic phosphate; interest; iterative design; lipophilicity; novel; polyanion; small molecule; stem; success; sugar; tool

Phosphorylated biomolecules play essential roles in human physiology, health, and medicine. Biological tar- gets for phosphorylation include nucleosides, lipids, amino acids, peptides, and proteins. It has been discovered recently that polyphosphorylation of proteins is an important post-translational modification, spurring researchers to synthesize chemical probes containing oligophosphate chains of specific lengths as tools to explore what has been termed the human polyP-ome. This development exposes the need for well-defined chemical reagents to enable phosphate chains of a desired length to be conjugated to an organic molecule of interest. Recently we reported the first well defined, crystalline reagent for the triphosphorylation of C, N, and O nucleophiles. This was obtained by activation of trimetaphosphate using a modern peptide coupling reagent, and now we propose to extend the methodology to afford new reagents for tetra-, penta-, and hexa-phosphorylation of C, N, and O nucle- ophiles; this is what we term our class I family of reagents for oligophosphorylation of organic molecules. We also propose to develop a class II family of reagents that is derived from the class I family by oligophosphorylation of the classic Wittig reagent, H2CPPh3. The class II family of reagents opens up the possibility to make the con- nection between an oligophosphate and a desired aldehyde-containing organic molecule via the Wittig reaction; in this case the constructs so obtained will contain a non-hydrolyzable P–C bond next to the new olefinic junction between the oligophosphate and the organic substrate. When using either the class I or II reagents to make the connection between an organic molecule and an oligophosphate chain, the initially formed product will contain an intact cyclophosphate residue. We propose to isolate and characterize such intermediates. Some of these will be stable under physiological conditions and will be targeted for further study. We will study the ring-opening of the cyclic intermediates by a variety of nucleophiles; use of hydroxide will give simply a terminal phosphate group at the end of the oligophosphate chain, while other nucleophiles are expected to result in target constructs that may contain linkages to two different organic residues at either end of the linear oligophosphate chain. In collabora- tion with the Raines group (MIT Chemistry) we propose to undertake collaborative biochemical studies of bovine pancreatic ribonuclease A (RNase A) inhibition by oligophosphates and their organic-molecule conjugates. Small molecule ribonuclease inhibitors are valuable biochemical tools for studies of RNA for which success often relies on shutting down all ribonucleolytic activity. Also in collaboration with Raines we propose to map out via protein crystallography the binding mode for oligophosphates and their organic conjugates in the RNase A active site, to aid in the iterative design of improved inhibitors.

磷酸化的生物分子在人体生理、健康和医学中发挥着重要作用。生物焦油-