Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1972 Apr;69(4):897–901. doi: 10.1073/pnas.69.4.897

Translational Restarts: AUG Reinitiation of a lac Repressor Fragment

Terry Platt 1, Klaus Weber 1, Don Ganem 1, Jeffrey H Miller 1,*
PMCID: PMC426590  PMID: 4554534

Abstract

An early, spontaneous amber mutation in the lac i-gene allows translational reinitiation, which results in a mutant lac repressor. Comparison of the amino-terminal sequence of this mutant repressor with the partial amino-acid sequence of the wild-type lac repressor shows that reinitiation occurs at the first internal AUG codon, and results in a mutant protein lacking 42 residues at the amino-terminal end. This protein binds the inducer isopropyl-β-D-thiogalactoside with normal affinity, and is capable of maintaining a tetrameric structure; however, it does not repress in vivo. These data suggest that the amino-terminal portion of the wild-type lac repressor is necessary either for direct binding to the lac operator or for the correct conformation for binding to DNA.

Keywords: protein biosynthesis, antibody, gel electrophoresis, amino-acid sequence, E. coli

Full text

PDF
897

Images in this article

899Image
on p.899
899Image
on p.899

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Capecchi M. R. Initiation of E. coli proteins. Proc Natl Acad Sci U S A. 1966 Jun;55(6):1517–1524. doi: 10.1073/pnas.55.6.1517. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Ghosh H. P., Söll D., Khorana H. G. Studies on polynucleotides. LXVII. Initiation of protein synthesis in vitro as studied by using ribopolynucleotides with repeating nucleotide sequences as messengers. J Mol Biol. 1967 Apr 28;25(2):275–298. doi: 10.1016/0022-2836(67)90142-8. [DOI] [PubMed] [Google Scholar]
  3. Gilbert W., Müller-Hill B. Isolation of the lac repressor. Proc Natl Acad Sci U S A. 1966 Dec;56(6):1891–1898. doi: 10.1073/pnas.56.6.1891. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Grodzicker T., Zipser D. A mutation which creates a new site for the re-initiation of polypeptide synthesis in the z gene of the lac operon of Escherichia coli. J Mol Biol. 1968 Dec;38(3):305–314. doi: 10.1016/0022-2836(68)90388-4. [DOI] [PubMed] [Google Scholar]
  5. Kumar S., Szybalski W. Orientation of transcription of the lac operon and its repressor gene in Escherichia coli. J Mol Biol. 1969 Feb 28;40(1):145–151. doi: 10.1016/0022-2836(69)90303-9. [DOI] [PubMed] [Google Scholar]
  6. Lengyel P., Söll D. Mechanism of protein biosynthesis. Bacteriol Rev. 1969 Jun;33(2):264–301. doi: 10.1128/br.33.2.264-301.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Michels C. A., Zipser D. Mapping of polypeptide reinitiation sites within the beta-galactosidase structural gene. J Mol Biol. 1969 May 14;41(3):341–347. doi: 10.1016/0022-2836(69)90280-0. [DOI] [PubMed] [Google Scholar]
  8. Miller J. H., Beckwith J., Muller-Hill B. Direction of transcription of a regulatory gene in E. coli. Nature. 1968 Dec 28;220(5174):1287–1290. doi: 10.1038/2201287a0. [DOI] [PubMed] [Google Scholar]
  9. Müller-Hill B., Crapo L., Gilbert W. Mutants that make more lac repressor. Proc Natl Acad Sci U S A. 1968 Apr;59(4):1259–1264. doi: 10.1073/pnas.59.4.1259. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Newton A. Effect of nonsense mutations on translation of the lactose operon of Escherichia coli. Cold Spring Harb Symp Quant Biol. 1966;31:181–187. doi: 10.1101/sqb.1966.031.01.026. [DOI] [PubMed] [Google Scholar]
  11. Newton A. Re-initiation of polypeptide synthesis and polarity in the lac operon of Escherichia coli. J Mol Biol. 1969 May 14;41(3):329–339. doi: 10.1016/0022-2836(69)90279-4. [DOI] [PubMed] [Google Scholar]
  12. Platt T., Miller J. H., Weber K. In vivo degradation of mutant lac repressor. Nature. 1970 Dec 19;228(5277):1154–1156. doi: 10.1038/2281154a0. [DOI] [PubMed] [Google Scholar]
  13. Sarabhai A., Brenner S. A mutant which reinitiates the polypeptide chain after chain termination. J Mol Biol. 1967 Jul 14;27(1):145–162. doi: 10.1016/0022-2836(67)90357-9. [DOI] [PubMed] [Google Scholar]
  14. Stewart J. W., Sherman F., Shipman N. A., Jackson M. Identification and mutational relocation of the AUG codon initiating translation of iso-1-cytochrome c in yeast. J Biol Chem. 1971 Dec 25;246(24):7429–7445. [PubMed] [Google Scholar]
  15. Weber K., Osborn M. The reliability of molecular weight determinations by dodecyl sulfate-polyacrylamide gel electrophoresis. J Biol Chem. 1969 Aug 25;244(16):4406–4412. [PubMed] [Google Scholar]
  16. Zipser D., Zabell S., Rothman J., Grodzicker T., Wenk M. Fine structure of the gradient of polarity in the z gene of the lac operon of Escherichia coli. J Mol Biol. 1970 Apr 14;49(1):251–254. doi: 10.1016/0022-2836(70)90392-x. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES