Analysis of translational fidelity of ribosomes with protamine messenger RNA as a template

Nozomu Mori, Yoshiko Funatsu, Kuniko Hiruta, S. Goto

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

A novel method was developed to estimate the translational fidelity of mammalian ribosomes in vitro with protamine mRNA of rainbow trout as template. Protamines are mixtures of basic proteins consisting of only seven types of amino acids (Arg, Ile, Val, Ser, Pro, Ala, and Gly), arginine (codon, AGR and CGN) being abundant. Taking advantage of the absence of lysine (codon, AAG) in the proteins, we determined the misincorporation of this amino acid into protamines in a cell-free translation system consisting of mouse liver ribosomes, protamine mRNA, [3H]lysine, [14C]arginine, and seven unlabeled amino acids: Ile, Val, Ser, Pro, Ala, Gly, and Met. After the reaction, translation products were analyzed by either sucrose gradient centrifugation or polyacrylamide gel electrophoresis. In the former method, radioactive protamines are mostly found on monosomes, but not on polysomes, probably because of the basic nature of the proteins. The error frequency was calculated from the molar ratio of [3H]lysine to [14C]arginine incorporated into protamines with an appropriate correction. The frequency was found to be 0.0006-0.002. This method enabled us to determine the frequency of misrecognition of purine bases at the second position of arginine codons in mRNA.

Original languageEnglish
Pages (from-to)1231-1239
Number of pages9
JournalBiochemistry
Volume24
Issue number5
Publication statusPublished - 1985

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Protamines
Ribosomes
Messenger RNA
Arginine
Codon
Lysine
Amino Acids
Proteins
Polyribosomes
Cell-Free System
Centrifugation
Oncorhynchus mykiss
Electrophoresis
Liver
Sucrose
Polyacrylamide Gel Electrophoresis

ASJC Scopus subject areas

  • Biochemistry

Cite this

Analysis of translational fidelity of ribosomes with protamine messenger RNA as a template. / Mori, Nozomu; Funatsu, Yoshiko; Hiruta, Kuniko; Goto, S.

In: Biochemistry, Vol. 24, No. 5, 1985, p. 1231-1239.

Research output: Contribution to journalArticle

Mori, Nozomu ; Funatsu, Yoshiko ; Hiruta, Kuniko ; Goto, S. / Analysis of translational fidelity of ribosomes with protamine messenger RNA as a template. In: Biochemistry. 1985 ; Vol. 24, No. 5. pp. 1231-1239.
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