Nishikimi M, Koshizaka T, Kondo K, Ozawa T, Yagi K
Department of Biomedical Chemistry, Faculty of Medicine, University of Nagoya, Japan.
Experientia. 1989 Feb 15;45(2):126-9. doi: 10.1007/BF01954844.
A mutant strain of Wistar rats with L-gulono-gamma-lactone oxidase deficiency has recently been established. To investigate this deficiency by DNA and RNA blot hybridization analyses, a fragment of a previously cloned cDNA encoding rat L-gulono-gamma-lactone oxidase was used as a probe. When genomic DNA of the mutant rat was digested with several restriction enzymes, the probe hybridized to fragments of the same sizes as those produced from DNA of normal rats. Poly(A)+RNA from the liver of the mutant rat was found to contain an L-gulono-gamma-lactone oxidase-specific mRNA of a normal size at a comparable level to that of normal rats. An in vitro translation experiment revealed that the mRNA programmed the synthesis of an enzyme protein which had the same molecular weight as that of the translational product of the normal mRNA, although the amount synthesized was markedly reduced as compared with that synthesized with the normal mRNA. In accordance with this observation, a very low but definite degree of L-gulono-gamma-lactone oxidase activity was detected in the microsomes of the mutant rat by a newly developed, highly sensitive method.
最近培育出了一种L-古洛糖酸-γ-内酯氧化酶缺乏的Wistar大鼠突变株。为了通过DNA和RNA印迹杂交分析来研究这种缺乏情况，一个先前克隆的编码大鼠L-古洛糖酸-γ-内酯氧化酶的cDNA片段被用作探针。当用几种限制性酶消化突变大鼠的基因组DNA时，该探针与从正常大鼠DNA产生的相同大小的片段杂交。发现来自突变大鼠肝脏的聚腺苷酸加尾RNA（Poly(A)+RNA）含有正常大小的L-古洛糖酸-γ-内酯氧化酶特异性mRNA，其水平与正常大鼠相当。体外翻译实验表明，该mRNA指导合成一种酶蛋白，其分子量与正常mRNA的翻译产物相同，尽管与用正常mRNA合成的量相比，合成量明显减少。根据这一观察结果，通过一种新开发的高灵敏度方法，在突变大鼠微粒体中检测到了非常低但确定程度的L-古洛糖酸-γ-内酯氧化酶活性。
