Purification and Properties of Double-stranded RNA-degrading Nuclease,dsRNase, from the Digestive Juice of the Silkworm, Bombyx mori
Purification and Properties of Double-stranded RNA-degrading Nuclease,dsRNase, from the Digestive Juice of the Silkworm, Bombyx mori
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DOI：
10.11416/jibs.76.1_57
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发表时间：
2007-02
影响因子：
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通讯作者：
Y. Arimatsu;Tetsuo Furuno;Y. Sugimura;M. Togoh;R. Ishihara;M. Tokizane;E. Kotani;Y. Hayashi;T. Furusawa
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文献类型：
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作者：
Y. Arimatsu;Tetsuo Furuno;Y. Sugimura;M. Togoh;R. Ishihara;M. Tokizane;E. Kotani;Y. Hayashi;T. Furusawa
碱性和酸性核酸酶，如核糖核酸酶和脱氧核糖核酸酶，已被研究与家蚕生长发育有关。一种酸性核糖核酸酶分布在家蚕的后部丝腺和脂肪体中，并在每龄蜕皮时活性增加，并与变态期间丝腺的组织分解有关(Koga等人，1969；Koga和Akune，1972)。在消化液和中肠组织中发现了一种依赖镁的碱性核酸酶，它能降解DNA和RNA(Mukai，1965；Funaguma et al.，1977)。此外，在蚕蛹中发现了一种依赖镁的糖非特异性核酸酶，它作用于天然和热变性的DNA和RNA(Mukai，1965)。核糖核酸酶III是从小牛胸腺(Ohtsuki等人，1977)、大肠杆菌(Robertson等人，1968；Young和Steitz，1978；Gotoh等人，1974)和鸡胚胎(Hall和Crouch，1977)中分离和纯化的双链RNA(DsRNA)。这种类型的核糖核酸酶似乎在处理特定的前体RNA以激活如tRNA、rRNA和mRNA的过程中发挥功能，并已被证明以一种循序渐进的方式消化dsRNA：特定切割的初始阶段，随后随机降解(Edy等人，1976)。家蚕的中肠和消化液已被发现具有核糖核酸酶活性，其降解dsRNA类似于合成的Poly(I)/Poly(C)和细胞质多角体病毒(CPV)的天然dsRNA基因组(Furusawa等人，1993)。这种酶暂时被称为家蚕dsRNase。然而，实际的酶特性仍有待澄清。本文报道了从家蚕消化液中成功分离出dsRNase，并对其底物专一性、辅因子需求和dsRNase活性的pH-活性曲线等酶性质进行了研究。
Alkaline and acid nucleases such as RNase and DNase have been studied with respect to the growth and development of the silkworm, Bombyx mori. An acid RNase is distributed in the posterior silk gland and fat body in B. mori, and shows an increase in activity at the ecdysis of each instar and in association with the histolysis of the silk gland during metamorphosis (Koga et al., 1969; Koga and Akune, 1972). A Mg -dependent alkaline nuclease, which degrades both DNA and RNA, was detected in the digestive juice and midgut tissues (Mukai, 1965; Funaguma et al., 1977). In addition, an Mg dependent sugarnon-specific nuclease which acts on both native and heatdenatured DNA and RNA has been found in pupae of the silkworm (Mukai, 1965). RNase III, specific for double-stranded RNA (dsRNA), has been isolated and purified from calf thymus (Ohtsuki et al., 1977), Escherichia coli (Robertson et al., 1968; Young and Steitz, 1978; Gotoh et al., 1974) and chick embryos (Hall and Crouch, 1977). This type of RNase appears to function in the processing of specific precursor RNA to activate such as tRNA, rRNA and mRNA, and has been shown to digest dsRNA in a step-wise manner: an initial phase of specific cleavage, followed by random degradation (Edy et al., 1976). The midgut and digestive juice of the silkworm have been found to possess an RNase activity, which degrades dsRNA similar to synthetic Poly(I)/Poly(C) and the natural dsRNA genome of the cytoplasmic polyhedrosis virus (CPV) (Furusawa et al., 1993). This enzyme was tentatively called Bombyx dsRNase. However, the actual enzymatic characterization remains to be clarified. In this paper, we describe successful isolation of dsRNase from digestive juice of the silkworm, and characterization of its enzymatic properties such as substrate specificity, co-factor requirement and pH-activity profile in dsRNase activity.