Enhanced synthetic luciferin substrates for firefly luciferase

萤火虫荧光素酶的增强型合成荧光素底物

• 批准号: 8532676
• 负责人: STEPHEN C. MILLER
• 金额: $ 34.9万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-26 至 2015-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8532676
• 关键词: Active Biological Transport; Adopted; Affinity; Animal Model; Binding; Biological; Biological Assay; Bioluminescence; Carbon Dioxide; Cells; Chemicals; Cyclization; Cytolysis; Detection; Disease Progression; Drug Efflux; Evaluation; Exhibits; Firefly Luciferases; Fluorescence; Gene Expression; Gene Expression Profiling; Image; In Vitro; Life; Light; Light Cell; Luciferases; Mammalian Cell; Methods; Mutation; Optical reporter; Organism; Output; Penetration; Permeability; Pharmaceutical Preparations; Photochemistry; Property; Pump; Reaction; Reporter Genes; Research; Rotation; Therapeutic; Tissues; Whole Organism; Work; base; bioluminescence imaging; cost; drug candidate; drug discovery; high throughput screening; imaging modality; improved; in vivo; inhibitor/antagonist; light emission; luciferin; mutant; novel therapeutics; protein protein interaction; quantum; response; screening; small molecule

Bioluminescence assays and imaging methods have been widely adopted for drug discovery because of their simplicity, robustness, and low cost. The inherently low background and lack of need for excitation light makes bioluminescence superior to fluorescence methods for many applications. Nevertheless, luciferase has yet to reach its full potential. Light emission from luciferase is limited by 1) access to the luciferin substrate (modulated by affinity, cell-permeability, and active transport by drug pumps), and 2) the photophysical properties of the luciferin (efficiency and wavelength of emission). To maximize the power of luciferase for biological applications we have synthesized new luciferin substrates that enhance luciferase light emission. We propose the following specific aims: 1) synthesis of new luciferin substrates with enhanced light emission properties; 2) identification of mutant luciferases that efficiently utilize synthetic luciferins; 3) evaluation of synthetic luciferins and mutant luciferases for live cell bioluminescence imaging.

生物发光测定和成像方法已被广泛采用 药物发现因其简单性、稳健性和低成本而受到关注。本来就低 背景和不需要激发光使得生物发光优于 荧光方法的许多应用。然而，荧光素酶尚未达到 它的全部潜力。荧光素酶的光发射受到以下因素的限制：1) 获取荧光素 底物（通过亲和力、细胞渗透性和药物泵的主动转运调节）， 2) 荧光素的光物理特性（效率和波长） 排放）。为了最大限度地发挥荧光素酶在生物应用中的作用，我们有 合成了增强荧光素酶发光的新荧光素底物。我们建议 具体目标如下：1）合成具有增强光作用的新型荧光素底物 发射特性； 2) 鉴定有效利用合成荧光素酶的突变体 荧光素； 3) 活细胞合成荧光素和突变荧光素酶的评价 生物发光成像。