Binding and catalytic properties of Xenopus (6-4) photolyase
Binding and catalytic properties of Xenopus (6-4) photolyase
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DOI：
10.1074/jbc.272.51.32591
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发表时间：
1997-12-19
影响因子：
4.8
通讯作者：
Todo, T
中科院分区：
文献类型：
--
作者：
Hitomi, K;Kim, ST;Todo, T
非洲爪蟾（6-4）光裂合酶与带有（6-4）光产物的DNA有高亲和力的结合并在光依赖性反应中修复它。为了阐明（6-4）光裂合酶的修复机制，我们用在单一位置带有光产物的合成DNA底物测定了它的结合和催化性质。（6-4）光裂合酶以高亲和力（K-D = 10（-9））结合双链DNA中的T[6 - 4]T，并结合单链DNA中的T[6-4]T和双链和单链DNA中的T[杜瓦]T，但亲和力略低（K-D =类似于2 x 10（-8））。大多数T[6-4]T-（6-4）光裂合酶复合物的解离非常缓慢（k（off）= 2.9 x 10（-5）s（-1））。在850-600 nm区域没有第二发色团的绝对作用光谱与酶的吸收光谱非常匹配。修复的量子产率（phi）约为0.11。（6-4）光裂合酶的完全还原形式（E-FADH（-））具有催化活性。对光活化产物的直接分析表明，（6-4）光裂合酶恢复了原始嘧啶。这些结果表明，顺，顺-环丁烷嘧啶二聚体光解酶和（6-4）光解酶是非常相似的，但它们的结合特性是不同的。
Xenopus (6-4) photolyase binds with high affinity to DNA bearing a (6-4) photoproduct and repairs it in a light-dependent reaction, To clarify its repair mechanism of (6-4) photolyase, we determined its binding and catalytic properties using synthetic DNA substrate which carries a photoproduct at a single location. The (6-4) photolyase binds to T[6-4]T in double-stranded DNA with high affinity (K-D = 10(-9)) and to T[6-4]T in single-stranded DNA and T[Dewar]T in double-and single-stranded DNA although with slightly lower affinity (K-D = similar to 2 x 10(-8)). Majority of the T[6-4]T-(6-4) photolyase complex dissociates very slowly (k(off) = 2.9 x 10(-5) s(-1)). Its absolute action spectrum without a second chromophore in the 850-600 nm region closely matches the absorption spectrum of the enzyme. The quantum yield (phi) of repair is approximately 0.11. The fully reduced form (E-FADH(-)) of (6-4) photolyase is catalytically active. Direct analysis of the photoreactivated product showed that (6-4) photolyase restores the original pyrimidines. These findings demonstrate that cis,syn-cyclobutane pyrimidine dimer photolyase and (6-4) photolyase are quite similar, but they are different with regard to the binding properties.