Tomarev S I, Zinovieva R D, Guo K, Piatigorsky J
Laboratory of Molecular and Developmental Biology, National Eye Institute, Bethesda, Maryland 20892.
J Biol Chem. 1993 Feb 25;268(6):4534-42.
Glutathione S-transferase (GST, EC 2.5.1.18) was purified from the digestive gland of the squid Ommastrephes sloani pacificus. It had high enzymatic activity for the 1-chloro-2,4-dinitrobenzene substrate and was composed of a major and a minor polypeptide band, both with molecular masses near 25 kDa on SDS-polyacrylamide gels. GST cDNA clones were derived from the digestive gland mRNA. The deduced GSTs of the longest cDNAs (pGST5 and pGST11) containing the entire coding sequence have a molecular mass near 23 kDa. Sequence comparisons showed that the squid GST is 42-44% identical to both squid and octopus S-crystallins (the major proteins of the lens), 32-34% identical to class pi and 29-32% identical to class alpha GSTs of vertebrates, and 19-23% identical to other GSTs of vertebrates and insects. Northern blot hybridization revealed that GST mRNAs were much more abundant in the digestive gland than in the testis, mantle, or lens. Analysis of a squid GST gene indicated that it has an exon-intron structure similar to that of the vertebrate class pi GST gene. An apparently novel repetitive element was identified in the 5'-flanking sequence of the squid GST gene. Our results suggest that multiple duplications of an ancestral GST gene gave rise to a family of enzymatically inactive crystallins specialized for lens refraction and one (or two) active GST enzyme expressed preferentially, but not exclusively, in the digestive gland in squids. This differs from the innovation of refractive function from a metabolic enzyme by increased expression in the lens with minimal or no gene duplication, as occurred among the enzyme-crystallins of vertebrates.
谷胱甘肽S-转移酶（GST，EC 2.5.1.18）从太平洋褶柔鱼的消化腺中纯化得到。它对1-氯-2,4-二硝基苯底物具有高酶活性，由一条主要多肽带和一条次要多肽带组成，在SDS-聚丙烯酰胺凝胶上两者分子量均接近25 kDa。GST cDNA克隆源自消化腺mRNA。包含完整编码序列的最长cDNA（pGST5和pGST11）推导的GST分子量接近23 kDa。序列比较表明，该鱿鱼GST与鱿鱼和章鱼的S-晶体蛋白（晶状体的主要蛋白质）有42-44%的同一性，与脊椎动物的π类GST有32-34%的同一性，与α类GST有29-32%的同一性，与脊椎动物和昆虫的其他GST有19-23%的同一性。Northern印迹杂交显示，GST mRNA在消化腺中的丰度远高于睾丸、外套膜或晶状体。对鱿鱼GST基因的分析表明，它具有与脊椎动物π类GST基因相似的外显子-内含子结构。在鱿鱼GST基因的5'侧翼序列中鉴定出一个明显新颖的重复元件。我们的结果表明，一个祖先GST基因的多次复制产生了一个专门用于晶状体折射的无酶活性的晶体蛋白家族，以及一种（或两种）主要在鱿鱼消化腺中优先（但非唯一）表达的活性GST酶。这与脊椎动物的酶-晶体蛋白中通过晶状体中表达增加且基因复制最少或无基因复制而从代谢酶产生折射功能的创新不同。
