TIME-RESOLVED MAGNETIC CIRCULAR DICHROISM

时间分辨磁圆二色性

• 批准号: 2900657
• 负责人: DAVID S. KLIGER
• 金额: $ 18.05万
• 依托单位: UNIVERSITY OF CALIFORNIA SANTA CRUZ
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-07-01 至 2000-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2900657
• 关键词: biomedical equipment development; chemical kinetics; circular magnetic dichroism; conformation; cytochrome b; cytochrome c; cytochrome oxidase; electron transport; enzyme mechanism; enzyme structure; hemoglobin; hemoprotein structure; magnetic field; method development; molecular dynamics; mutant; photochemistry; photolysis; protein structure function; structural biology; time resolved data; ultraviolet radiation

DESCRIPTION: Development and use of time resolved magnetic circular dichroism spectroscopy (TRMCD) is proposed. The application of a magnetic field induces chirality and the observed signal is generally localized to the moiety who's absorption is being probed. This technique is used for heme centers. Extension of the technique to the near UV is to be developed, and preliminary data on the static MCD of tryptophan is encouraging (will require an extra electromagnet to compensate for the Faraday rotation of the cell windows). The central idea is to photolyze a ligand and then study the kinetics. The resolution of the TRMCD apparatus is about 50-100 ns in the heme center absorption (partly complicated by signals that result from the protein diffusional reorientation time, about 100 ns- this complication has been analyzed and is probably correctable). Studies on cytochrome oxidase, cytochrome c3, and cytochrome c' are ongoing. A particularly important finding of the past period was the finding of a transient ligation at the cytochrome a3 site of cytochrome oxidase, resulting in a model for part of the gating mechanism coupling energy release by oxygen reduction to proton pumping (the ligand shuttle model). An unknown ligand in the protein binds to the protein iron center and this has been best studied with this technique. Most interesting future studies center on hemoglobin and the R to T allosteric transition, if the trp residues can be isolated and measured. Some hope has been demonstrated by static MCD on trp in solution. Mutants should isolate which trp residues yield signal. Recent TRCD results, just submitted to Biochemistry, are exciting. Outstanding studies except there were two problems cited in last review: the problem of protein orientational relaxation, which is on the 100 ns time scale, and the issue of productivity.

描述：时间分辨磁环的开发和使用 提出了二色性光谱（TRMCD）。 的应用 磁场引起手性，观察到的信号通常是 定位于正在探测吸收的部分。 这 技术用于血红素中心。将该技术扩展到 近紫外区待开发，静态 MCD 的初步数据 色氨酸是令人鼓舞的（需要额外的电磁体 补偿单元窗口的法拉第旋转）。 中央 想法是光解配体，然后研究动力学。 这 TRMCD 装置在血红素中心的分辨率约为 50-100 ns 吸收（部分因蛋白质产生的信号而复杂化） 扩散重定向时间，大约 100 ns——这种并发症已被 已分析并且可能是可纠正的）。 细胞色素氧化酶的研究， 细胞色素 c3 和细胞色素 c' 正在进行中。 一个特别重要的 过去一段时间的发现是在 细胞色素氧化酶的细胞色素 a3 位点，产生部分模型 通过氧还原耦合能量释放的门控机制 质子泵浦（配体穿梭模型）。未知配体 蛋白质与蛋白质铁中心结合，这已得到充分研究 用这种技术。 未来最有趣的研究集中在 血红蛋白和 R 到 T 变构转变，如果色氨酸残基可以 被隔离和测量。静态 MCD 已经证明了一些希望 溶液中的色氨酸。突变体应分离产生哪些色氨酸残基 信号。最近刚刚提交给生物化学的 TRCD 结果是 令人兴奋。杰出的研究，除了其中引用的两个问题 上一篇回顾：蛋白质取向弛豫问题，即 在 100 ns 时间尺度上，以及生产力问题。