Haugen T H, Heath E C
Proc Natl Acad Sci U S A. 1979 Jun;76(6):2689-93. doi: 10.1073/pnas.76.6.2689.
The de novo biosynthesis of RNase (ribonucleate 3'-pyrimidino-oligonucleotidohydrolase, EC 3.1.4.22) was studied in a cell-free rabbit reticulocyte translation system using a polyadenylylated fraction of mRNA isolated from bovine pancreas. Analysis of the [35S]methionine-labeled translation products of pancreas mRNA by polyacrylamide gel electrophoresis indicated the presence of several proteins, one of which corresponded to 16,500 daltons, or approximately 2800 daltons greater than native RNase A. This protein was specifically precipitated from the mixture of products by anti-RNase antibodies. Partial sequence determination of the NH2-terminal region of the anti-RNase antibody-precipitable species indicated that it is a precursor form of RNase A with 25 additional amino acids on its NH2 terminus. The precursor nature of the protein was confirmed by demonstration that a mixture of RNase A and a glycosylated form of the enzyme, RNase B, is formed when translation of the mRNA is conducted in the presence of dog pancreas membranes. Assay of the putative precursor form of RNase for catalytic activity with polycytidylic acid as substrate indicated that the protein has a specific enzymatic activity identical to that of native RNase A.
利用从牛胰腺中分离出的聚腺苷酸化mRNA组分，在无细胞兔网织红细胞翻译系统中研究了核糖核酸酶（核糖核酸3'-嘧啶寡核苷酸水解酶，EC 3.1.4.22）的从头生物合成。通过聚丙烯酰胺凝胶电泳分析胰腺mRNA的[35S]甲硫氨酸标记的翻译产物，结果表明存在几种蛋白质，其中一种蛋白质的分子量为16,500道尔顿，比天然核糖核酸酶A大约大2800道尔顿。抗核糖核酸酶抗体可从产物混合物中特异性沉淀出这种蛋白质。对可被抗核糖核酸酶抗体沉淀的蛋白质的NH2末端区域进行部分序列测定，结果表明它是核糖核酸酶A的前体形式，其NH2末端有25个额外的氨基酸。当在犬胰腺膜存在的情况下进行mRNA的翻译时，会形成核糖核酸酶A和该酶的糖基化形式核糖核酸酶B的混合物，这证实了该蛋白质的前体性质。以聚胞苷酸为底物测定假定的核糖核酸酶前体形式的催化活性，结果表明该蛋白质具有与天然核糖核酸酶A相同的比酶活性。
