SYNTHETIC FLAVO HEMO PROTEINS

合成黄素血蛋白

• 批准号: 6480838
• 负责人: PETER LESLIE DUTTON
• 金额: $ 15.58万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-08-01 至 2002-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6480838
• 关键词: biological products; biomedical resource; proteins; technology /technique

We have succeeded in designing and constructing de novo synthetic flavo-hemo-proteins that exhibit light activated intraprotein electron transfer. We use our traditional four-helix bundle framework to covalently link a bromoflavin to an interior cysteine very close to the geometric center of the bundle. In the same bundle, interior histidines a positioned to form a bis-His ligation of added heme toward either end of the bundle interior. We have shown that illumination of the blue flavin absorption band creates an excited state which can extract a pair of electrons from a convenient nitrogenous base donor (such as EDTA) to create a reduced flavin that spontaneously transfers electrons to nearby bis-histidine ligated heme. We are presently using the RLBL nanosecond dye laser to capture the spectra of the excited flavin state, the subsequent reduced flavin, and to provide an initial description of the time course of intraprotein electron transfer from the reduced fla vin to the heme. It is quite possible that there will be a heterogeneity of the time course corresponding to a distribution of flavin/heme distances for different conformations of the redox peptide. We expect a competition between flavin excited state quenching by energy transfer to the heme and electron transfer between the flavin and the heme. We will be flowing the synthetic flavo-hemo-protein plus EDTA solution through a cell to compare the spectra of regions with and without the blue pump pulse. We can easily regenerate our material by exposing the solution to molecular oxygen, which rapidly oxidizes the heme component. In addition, we are investigating the viability of using a tryptophan moiety in this electron transfer scheme. The tryptophan residue will be pumped with the fourth harmonic of the Nd:YAG (266 nm).

我们已经成功地设计和构建了从头合成的 显示光激活蛋白内电子的黄素血蛋白 转移 我们使用传统的四螺旋束框架， 将溴黄素与内部半胱氨酸共价连接， 束的几何中心。 在同一束，内部 组氨酸被定位以形成附加血红素的双His连接 朝向束内部的任一端。 我们已经证明 蓝色黄素吸收带的照明产生了一个激发的 一种可以从一个方便的电子中提取一对电子的状态， 含氮碱供体（如EDTA）以产生还原的黄素， 自发转移电子到附近的双组氨酸连接 血红素 我们目前正在使用RLBL纳秒染料激光器来捕获 激发黄素态的光谱，随后的还原 黄素，并提供一个初步的时间过程的描述， 从还原的弗拉vin到血红素的蛋白内电子转移。 很有可能会出现时间的异质性 对应于黄素/血红素距离分布的过程， 氧化还原肽的不同构象。 我们期待一场比赛 黄素激发态通过能量转移到血红素而淬灭 以及黄素和血红素之间的电子转移。 我们将 使合成的黄素血红蛋白加EDTA溶液流过 单元，用于比较具有和不具有蓝色泵浦的区域的光谱 脉搏了 我们可以很容易地再生我们的材料， 分子氧迅速氧化血红素成分。 在 此外，我们正在研究使用色氨酸的可行性 在这个电子转移方案中的部分。 色氨酸残基会 用Nd：YAG的四次谐波（266 nm）泵浦。