ABSOLUTE ACTION SPECTRUM OF E-FADH2 AND E-FADH2-MTHF FORMS OF ESCHERICHIA-COLI DNA PHOTOLYASE
ABSOLUTE ACTION SPECTRUM OF E-FADH2 AND E-FADH2-MTHF FORMS OF ESCHERICHIA-COLI DNA PHOTOLYASE
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DOI：
10.1021/bi00485a021
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发表时间：
1990-08-21
期刊：
影响因子：
2.9
通讯作者：
SANCAR, A
中科院分区：
文献类型：
--
作者：
PAYNE, G;SANCAR, A
大肠杆菌DNA光解酶介导紫外线损伤DNA中嘧啶二聚体的光修复。该酶含有两个发色团，1，5-二氢黄素腺嘌呤二核苷酸（FADH 2）和5，10-亚甲基四氢叶酸聚谷氨酸（MTHF）。为了确定两个发色团在光化学反应中导致DNA修复的作用以及DNA结构对光催化步骤的影响，我们确定了仅含FADH 2（E-FADH 2）或两个发色团（E-FADH 2-MTHF）的酶的绝对作用光谱，双链和单链底物以及胸腺嘧啶二聚体附近不同序列的底物。我们发现E-FADH 2的作用光谱的形状与具有量子产率vphi的吸收光谱的形状相匹配。（FADH 2）= 0.69。E-FADH 2-MTHF的作用光谱也与vphi的吸收光谱相当好地一致。（FADH 2-MTHF）= 0.59。从这些值和从先前建立的两种发色团的性质，我们提出MTHF以v φ的量子产率将能量转移到FADH 2。T = 0.8，并且1FADH 2单线态以量子产率v φ ET = 0.69将电子转移到二聚体或从二聚体转移电子。发色团的化学性质在几个催化循环后没有改变。该酶修复胸腺嘧啶二聚体在五个不同的序列背景下具有相同的效率。类似地，单链和双链DNA以相同的总量子产率被修复。
Escherichia coli DNA photolyase mediates photorepair of pyrimidine dimers occurring in UV-damaged DNA. The enzyme contains two chromophores, 1,5-dihydroflavin adenine dinucleotide (FADH2) and 5,10-methenyltetrahydrofolylpolyglutamate (MTHF). To define the roles of the two chromophores in the photochemical reaction(s) resulting in DNA repair and the effect of DNA structure on the photocatalytic step, we determined the absolute action spectra of the enzyme containing only FADH2 (E-FADH2) or both chromophores (E-FADH2-MTHF), with double- and single-stranded substrates and with substrates of different sequences in the immediate vicinity of the thymine dimer. We found that the shape of the action spectrum of E-FADH2 matches that of the absorption spectrum with a quantum yield .vphi.(FADH2) = 0.69. The action spectrum of E-FADH2-MTHF is also in a fairly good agreement with the absorption spectrum with .vphi.(FADH2-MTHF) = 0.59. From these values and from the previously established properties of the two chromophores, we propose that MTHF transfers energy to FADH2 with a quantum yield of .vphi..epsilon.T = 0.8 and that 1FADH2 singlet transfers an electron to or from the dimer with a quantum yield .vphi.ET = 0.69. The chemical nature of the chromophores did not change after several catalytic cycles. The enzyme repaired a thymine dimer in five different sequence contexts with the same efficiency. Similarly, single- and double-stranded DNAs were repaired with the same overall quantum yield.