Biochemical characteristics of cyclobutane pyrimidine dimer photolyase and UV-B protecting mechanisms of mice (Oryza sativa L.)

我们在2000年和2002年期间有几项新发现：光敏nice Norin 1缺乏环丁烷嘧啶（CPDs）的光修复。利用闪光灯的方法，我们发现体内光修复的缺陷是由光分解酶的结构/功能改变引起的。这些结果被提取物的研究证实，表明Norin 1光解酶二聚体复合物相对于抗紫外线的Sasanishiki光解酶具有较高的耐热性。2：克隆了Sasanishiki和Norin 1的CPD光解酶基因并进行了测序。他们被发现编码506个氨基酸残基的ORF。第126位的氨基酸在Sasanishiki中为谷氨酰胺，而在Norin 1.3中为精氨酸。从大肠杆菌KY20中纯化出gst融合水稻CPD光解酶，我们确定FAD是两个发色团之一。4：深色生长的黄化水稻幼苗几乎没有CPD光解酶活性。光解酶基因的表达受光敏色素和/或隐色素的光控制。5：我们开始用农杆菌转化系统将CPD光解酶基因转化为转基因水稻植株。6 .遗传背景为台中65 （T-65）的水稻紫色叶基因pl近等基因系（NIL）在UV-B升高的培养过程中，尽管吸收uv的化合物和花青素的积累比T-65高，但其生长明显低于T-65。NIL的稳态CPD水平也高于T-65。我们发现，在可见光照射下，慢性UV-B照射下的生长发育迟缓和CPD水平升高可能是由于有效产生CPD水平的UV-B和有效抑制CPD光解酶活性的蓝/UV-A的筛选作用不同。

We made several new findings in the period of 2000 and 2002.1:UV-sensitive nice Norin 1 is deficient in photorepair of cyclobutane pyrimidine diners (CPDs). Using a photoflash approach, we showed that the deficiency in photorepair in vivo resulted from a structure/function alteration photolyase. These results were confirmed by studies with extracts, which showed that the Norin 1 photolyase-dimer complex was highly thermolabile relative to the UV-resistant Sasanishiki photolyase.2:CPD photolyase gene from Sasanishiki and Norin 1 were cloned and sequenced. They were found to encode an ORF for 506 amino acid residues. Amino acid at position 126 was glutamine in Sasanishiki, but was arginine in Norin 1.3:GST-fusion rice CPD photolyase was purified from E. coli KY20, and we determined FAD as one of the two chromophores.4:Dark-grown etiolated rice seedlings did hardly have CPD photolyase activity. The induction of CPD photolyase in dark-grown seedlings was under photocontrol of phytochrome, and the expression of photolyase gene was under photocontrol of phytochrome and/or cryptochrome.5:We started to produce a transgenic rice plant into which CPD photolyase gene was transformed using the Agrobacterium transformation system.6:The growth of near-isogenic line (NIL) of rice for purple leaf gene pl of rice with a genetic background of Taichung 65 (T-65) was significantly decreased relative to T-65 in the elevated UV-B during culture regardless of higher accumulation of UV-absorbing compounds and anthocyanins than T-65. The steady-state CPD levels in the NIL was also higher than that in T-65. We showed that the growth stunting and increase in CPD levels under chronic UV-B irradiated with visible light might be due to the difference in the screening effects of UV-B effective for producing CPD levels and blue/UV-A effective for CPD photolyase activity.