Polypetide chain termination. Purification of the release factors, R1 and R2, from Escherichia coli.

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Publication Type Journal Article
School or College College of Science; School of Medicine
Department Oncological Sciences; Biology; Human Genetics
Program Institute of Human Genetics; Howard Hughes Medical Institute (HHMI)
Creator Capecchi, Mario R.
Other Author Klein, Herbert A.
Title Polypetide chain termination. Purification of the release factors, R1 and R2, from Escherichia coli.
Date 1971-02-25
Description We have extensively purified the release factors RI and Rz from Escherichia coli. These proteins can each mediate polypeptide chain termination. The physiological substrate for this reaction is a completed polypeptide chain in a peptidyl- transfer RNA-messenger RNA-ribosome complex. The reaction consists of recognition of a chain-terminating signal in the mRNA and hydrolysis of the peptidyl-tRNA ester bond, releasing the polypeptide chain. For the purification we were guided by two kinds of assay, the hexapeptide release assay and the formylmethionme release assay, each named according to the molecule released in a model reaction analogous to physiological chain termination. The two factors have different codon specificities, RI acting in response to UAG or UAA, and Re in response to UGA or UAA. Both factors were purified from a l-kg batch of frozen E. coli MRE600 by a scheme which carried the material through four steps before reaching a branch point at the Bth step, when RI- and R2-rich fractions were produced. The two fractions were then treated similarly but separately through three more steps. The products were studied by polyacrylamide gel lectrophoresis using both routine and sodium dodecyl sulfate techniques. For the 6.4 mg of purified RI, we estimate 85% purity after 2,000-fold purification. For the 9.4 mg of purified Rz, we estimate 99% purity after 1,500-fold purification. Our results indicate that each release factor consists of a single polypeptide chain with a molecular weight of 44,000 for RI and 47,000 for Rz. We calculate that there are about 500 molecules of RI and 700 molecules of Rz per E. coli cell.
Type Text
Publisher American Society for Biochemistry and Molecular Biology (ASBMB)
Volume 246
Issue 4
First Page 1055
Last Page 1061
Subject Acrylates; Detergents
Subject MESH Bacterial Proteins
Language eng
Bibliographic Citation J Biol Chem. 1971 Feb 25;246(4):1055-61: Polypetide chain termination. Purification of the release factors, R1 and R2, from Escherichia coli. Klein H, Capecchi MR. Retrieved on September 22.2006 from http://www.jbc.org/cgi/reprint/246/4/1055.
Rights Management Copyright © 1971 American Society for Biochemistry and Molecular Biology. All Rights Reserved..
Format Medium application/pdf
Identifier ir-main,456
ARK ark:/87278/s60g43m5
Setname ir_uspace
Date Created 2012-06-13
Date Modified 2021-05-06
ID 705806
Reference URL https://collections.lib.utah.edu/ark:/87278/s60g43m5
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