中文摘要
本项目运用短脉冲电子束、激光束与太赫兹时域光谱技术，结合稳态产物分析及酶活性分析，从微观、动态和分子反应的角度上研究了自由基、光敏剂与对溶菌酶、细胞色素P450酶等蛋白质的电荷与能量转移机理和动力学过程，结果表明在有氧条件下核黄素对溶菌酶的损伤是Ⅰ型和Ⅱ型反应协同作用的结果, 并以Ⅱ型反应为主。荧光分析实验表明Trp 62是核黄素诱导溶菌酶氧化损伤的位点之一；激光光解研究表明，在355 nm激光作用下核黄素能够通过电子转移反应选择性损伤特定氨基酸如色氨酸和酪氨酸，形成的氨基酸自由基之间能够成键，导致溶菌酶发生双分子交联。核黄素与细胞色素P450之间电子转移反应的研究表明，核黄素阴离子自由基可以迅速地质子化而以核黄素中性自由基的形式存在，其可以还原CYP450辅基亚铁血红素上的三价铁生成还原态的CYP450。 本项目还利用THz-TDS技术进行了相关生物分子的结构及定量分析研究，为今后进一步的研究打下了良好基础。
英文摘要
Proteins are the major target for reactive oxygen species (ROS) as a result of their abundance in biological systems.In this work the property of photo-sensitizer and its reaction with protein were studied using steady state study together with time-resolved technology. The photooxidation damage of lysozyme in the presence of riboflavin was studied and results showed both type I and type II process were observed in the present of oxgygen and type II was the dominant process. Fluorescence spectroscopy indicated that one of the damaged sites on lysozme was Trp 62. The results of transient study showed that lysozyme was oxidative damaged via electron thranfer reaction from triplet riboflavin under nitrogen. The study also suggested that antioxidants, such as epicatechin, protected lysozyme via quenching triplet riboflavin effectively. The electron transfer reaction between riboflavin and cytochrome P450 in rat liver microsomes was studied and it showed that anion radical of riboflavin chould be protonate to be RFH? which can reduce the heme iron forming reductant cytochrome P450. And Terahertz time–domain spectroscopy (THz–TDS) was applied for quantitatively analysing a series of binary mixtures and a ternary mixture too.
结题摘要
本项目运用短脉冲电子束、激光束与太赫兹时域光谱技术，结合稳态产物分析及酶活性分析，从微观、动态和分子反应的角度上研究了自由基、光敏剂与对溶菌酶、细胞色素P450酶等蛋白质的电荷与能量转移机理和动力学过程，结果表明在有氧条件下核黄素对溶菌酶的损伤是Ⅰ型和Ⅱ型反应协同作用的结果, 并以Ⅱ型反应为主。荧光分析实验表明Trp 62是核黄素诱导溶菌酶氧化损伤的位点之一；激光光解研究表明，在355 nm激光作用下核黄素能够通过电子转移反应选择性损伤特定氨基酸如色氨酸和酪氨酸，形成的氨基酸自由基之间能够成键，导致溶菌酶发生双分子交联。核黄素与细胞色素P450之间电子转移反应的研究表明，核黄素阴离子自由基可以迅速地质子化而以核黄素中性自由基的形式存在，其可以还原CYP450辅基亚铁血红素上的三价铁生成还原态的CYP450。 本项目还利用THz-TDS技术进行了相关生物分子的结构及定量分析研究，为今后进一步的研究打下了良好基础。