FLUORESCENCE ENERGY TRANSFER STUDIES OF PROTEIN INDUCED DNA BENDING

蛋白质诱导 DNA 弯曲的荧光能量转移研究

• 批准号: 6281092
• 负责人: R H EBRIGHT
• 金额: $ 2.13万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-08-01 至 1999-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6281092
• 关键词: biological products; biomedical resource; proteins

We have used time-resolved fluorescence energy transfer to investigate protein-induced DNA bending in solution. We are measuring multiple protein-to-DNA and DNA-to-DNA distances in each of two protein-DNA complexes that exhibit dramatic protein-induced DNA bending in the crystalline state: the CAP-DNA complex and the TBP-DNA complex. We are determining DNA bend angles and trajectories in solution, assessing the salt-dependence of the protein-induced DNA bending, assessing the effects of protein mutants on protein-induced DNA bending and assessing the effects of DNA mutants on protein-induced DNA bending. We are using AEDANS ??ex=340nm; ?em=480nm) as the donor fluorochrome and fluorescein (?ex=490 nm; ?em=520 nm) as the acceptor fluorochrome. We have incorporated AEDANS at specific sites in protein by site-directed mutagenesis to introduce unique surface cysteines followed by cysteine-specific chemical modification, and we have incorporated fluorescein at specific sites in DNA by total synthesis. Preliminary results have been reported.

我们用时间分辨荧光能量转移