Payne G, Heelis P F, Rohrs B R, Sancar A
Department of Biochemistry, School of Medicine, University of North Carolina, Chapel Hill 27514.
Biochemistry. 1987 Nov 3;26(22):7121-7. doi: 10.1021/bi00396a038.
Escherichia coli DNA photolyase is a flavoprotein that when purified is blue in color and contains a stable neutral radical FAD (E-FADH). In the presence of a suitable electron donor (i.e., thiols, tyrosine, or NADH) the radical FAD adsorbs visible light and undergoes photoreduction to the fully reduced FAD (E-FADH2). The in vitro quantum yield of dimer repair for E-FADH is 0.07 while that of E-FADH2 approaches the in vivo value of 1. Electron paramagnetic resonance studies on whole cells indicate that the in vivo form of photolyase is E-FADH2 with enzyme containing radical FAD generated predominantly during the ammonium sulfate precipitation step of the purification. Activity measurements of E-FADH using long-wavelength photoreactivating light indicate that enzyme containing FAD in the radical form is not active in dimer repair. Dimer repair observed with E-FADH at shorter wavelengths is probably photoreduction of E-FADH followed by dimer repair by E-FADH2.
大肠杆菌DNA光解酶是一种黄素蛋白，纯化后呈蓝色，含有稳定的中性自由基FAD（E-FADH）。在合适的电子供体（即硫醇、酪氨酸或NADH）存在下，自由基FAD吸收可见光并发生光还原，生成完全还原的FAD（E-FADH2）。E-FADH的二聚体修复体外量子产率为0.07，而E-FADH2的量子产率接近体内值1。对全细胞的电子顺磁共振研究表明，光解酶的体内形式是E-FADH2，含有自由基FAD的酶主要在纯化的硫酸铵沉淀步骤中产生。使用长波长光复活光对E-FADH进行活性测量表明，含有自由基形式FAD的酶在二聚体修复中无活性。在较短波长下用E-FADH观察到的二聚体修复可能是E-FADH的光还原，随后由E-FADH2进行二聚体修复。
