酿酒酵母中一种新型聚（U）、聚（C）核糖核酸酶的纯化与鉴定

Purification and characterization of a novel poly(U), poly(C) ribonuclease from Saccharomyces cerevisiae.

作者信息

Lalioti V S, Ballesta J P, Fragoulis E G

机构信息

University of Athens, Department of Biochemistry and Molecular Biology, Greece.

出版信息

Biochim Biophys Acta. 1997 Sep 26;1342(1):62-72. doi: 10.1016/s0167-4838(97)00078-2.

DOI:10.1016/s0167-4838(97)00078-2

PMID:9366271

Abstract

A new ribonuclease from Saccharomyces cerevisiae, specific for poly(U) and poly(C) substrate, was purified near to homogeneity by successive fractionation with DEAE-Sepharose, Heparin-Sepharose and CM-Sepharose chromatography. The purified molecule detected by SDS/polyacrylimide gel electrophoresis has a molecular mass of 29 kDa. The optimum pH for the enzyme activity is 5.5-7 and its isoelectric point is 7.5. The purified enzyme was able to degrade 26S, 18S and 5S rRNAs as well as mRNA obtained from in vitro transcription. No catalytic activity was observed when the RNase was incubated with tRNA and double stranded substrate. Our findings suggest that this novel RNase may play an important role in the processing of RNA in Saccharomyces cerevisiae.

摘要

从酿酒酵母中分离出一种新的核糖核酸酶，它对聚（U）和聚（C）底物具有特异性，通过用DEAE-琼脂糖、肝素-琼脂糖和CM-琼脂糖色谱进行连续分级分离，纯化至接近均一。通过SDS/聚丙烯酰胺凝胶电泳检测到的纯化分子的分子量为29 kDa。该酶活性的最适pH为5.5-7，其等电点为7.5。纯化后的酶能够降解26S、18S和5S rRNA以及体外转录获得的mRNA。当该核糖核酸酶与tRNA和双链底物一起孵育时，未观察到催化活性。我们的研究结果表明，这种新型核糖核酸酶可能在酿酒酵母的RNA加工过程中发挥重要作用。