The ribosome as 'affinity matrix': Efficient purification scheme for translation factors

被引：11

作者：

Ganoza, MC ^{[1
]}

Aoki, H ^{[1
]}

Burkhardt, N ^{[1
]}

Murphy, BJ ^{[1
]}

机构：

[1] MAX PLANCK INST MOLEC GENET, D-14195 BERLIN, GERMANY

来源：

BIOCHIMIE | 1996年 / 78卷 / 01期

关键词：

translation initiation; translation elongation factors;

D O I：

10.1016/0300-9084(96)81329-0

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

A convenient method to purify each of the non-ribosomal proteins required to translate a native mRNA in vitro is described. In this scheme, the ribosome is used as an 'affinity' matrix to selectively elute the non-ribosomal proteins required for translation that are bound to these particles. Different sets of these proteins can be eluted with solutions of Mg2+ and NH;: of various concentrations from either 70S, or 30S and 50S particles. A scheme for the purification of each initiation, elongation and release factor and 20 aminoacyl-tRNA synthetases is described. Specific examples of the purification of the initiation (IF-1, IF-2, IF-3) and elongation (EF-Tu and EF-G) factors and for a protein called 'rescue', which affects the association of native ribosomal subunits, are given. A scheme for the purification of EF-P, which stimulates peptide-bond synthesis and one of the W proteins, which permit reconstitution of translation is also described. The procedure markedly simplifies the isolation, in homogeneous form, of all the non-ribosomal proteins required to reconstruct translation.

引用

页码：51 / 61

页数：11

共 42 条

[1] CLONING, SEQUENCING AND OVEREXPRESSION OF THE GENE FOR PROKARYOTIC FACTOR-EF-P INVOLVED IN PEPTIDE-BOND SYNTHESIS [J].