Photobiological significance of DNA photolgase/Blue-light photoreceptor family

我们报道了非洲爪哇(6-4)光解酶基因的分离，并证明了(6-4)光解酶与化学计量比的FAD非共价结合。非洲爪哇(6-4)光解酶与带有(6-4)光产物的DNA高亲和力结合并在光依赖的反应中修复它。为了阐明其对(6-4)光解酶的修复机制，我们利用合成的DNA底物在单一位置携带光解产物来测定其结合和催化性质。(6-4)光解酶与双链DNA中的T[6-4]T有较高的亲和力(Kd=10~(-9)&gt；)，在单链DNA中与T[6-4]T结合，在双链和单链DNA中与T[Dewar]T结合，但亲和力稍低(k_D=-2×10~(-4))。大多数T[6-4]T(6-4)光解酶复合体的解离速度很慢(koff=2.9x10~(-5)S~(-1))。它的绝对作用光谱在350-600 nm范围内没有第二个生色团，与酶的吸收光谱非常匹配。修复的量子产额(F)约为0.11。…此外，(6-4)光解酶的完全还原形式(E-FADH-)具有催化活性。对光复活产物的直接分析表明，(6-4)光解酶恢复了原来的嘧啶类化合物。这些发现表明，CPD光解酶和(6-4)光解酶非常相似，但它们的结合性质不同。光是携带昼夜节律的主要环境信号。光信号可以通过改变它们的相位来改变这些节奏。然而，人们对光信号感知和转导的分子机制知之甚少。光解酶/隐花色素家族的成员含有黄素腺嘌呤二核苷酸(FAD)作为发色团，参与DNA修复和环境光信号的光接收两个不同的功能。我们报道了从果蝇中克隆出这个家族的一个新成员--Dcay。Northern印迹分析表明，该基因在多种组织中均有表达。。在成体头部中，dcrip mRNA以昼夜节律的方式表达，而在时钟缺陷的per0和tim0突变体中，这种节律波动被取消。昼夜节律的表达在持续的黑暗中变得黯淡。该基因的过度表达改变了果蝇运动节律中光诱导的相位延迟。这些结果表明，DCRY是一种昼夜节律光感受器，其表达受生物钟基因的调控。较少

We report isolation of a Xenopus laevis (6-4)photolyase gene, and show that the (6-4)photolyase binds noncovalently to stoichiometric amounts of FAD.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 (kD = 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 = -2 x 10_<-8>). Majority of the T[6-4]T (6-4) photolyase-complex dissociates very slowly (koff = 2.9 x 10_<-5 S-1>). Its absolute action spectrum without a second chromophore in the 350-600 nm region closely matches the absorption spectrum of the enzyme. The quantum yield (f) of repair is approximately 0.11. … More 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 CPD photolyase and (6-4)photolyase are quite similar, but they are different with regard to the binding properties.Light is the major environmental signal for the entrainment of circadian rhythms. Light signals can entrain these rhythms by shifting their phases. However, little is known about the molecular mechanism for the perception and transduction of the light signal. The members of the photolyase/cryptochrome family contain flavin adenine dinucleotide (FAD) as chromophore and are involved in two diverse functions, DNA repair and photoreception of environmental light signals. We report the cloning of a new member of this family, dcry, from Drosophila. Northern blot analysis shows that this gene is expressed in various tissues. . The dcry mRNA is expressed in a circadian manner in adult heads, while such rhythmic fluctuation is abolished in the clock-defective per0 and tim0 mutants. The circadian expression damps in constant darkness. Over-expression of the dcry gene alters the light-induced phase delay in the locomotor activity rhythms of flies. These results suggest that DCRY is a circadian photoreceptor and its expression is regulated by circadian clock genes.. Less