Skip to main content
NCBI 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 Feb;69(2):412–416. doi: 10.1073/pnas.69.2.412

β-Galactosidase: Immunological Activity of Ribosome-Bound, Growing Polypeptide Chains

Joyce Hamlin 1,*, Irving Zabin 1
PMCID: PMC426469  PMID: 4551143

Abstract

Ribosomes carrying nascent chains of β-galactosidase were prepared by disruption of Escherichia coli in detergent-free buffer of high salt concentration, followed by purification on a discontinuous sucrose gradient. Assay by the method of immune hemolysis inhibition with anti-β-galactosidase indicated that considerable amounts of antibody were bound by the growing chains. Much of the crossreacting material could be released from the ribosomes by treatment with puromycin. The ability to bind anti-β-galactosidase was completely destroyed when ribosomes were heated at 60°C. At very early times after induction, well before the appearance of active enzyme, crossreacting material could be demonstrated on ribosomes; this finding correlated with the appearance of an amino-terminal fragment of β-galactosidase. Thus, growing chains of β-galactosidase must begin to fold before their release from the ribosome.

Keywords: immune hemolysis inhibition, antiserum, puromycin, protein conformation, chain folding

Full text

PDF
412

Selected References

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

  1. ARQUILLA E. R., HAMASHIGE S., HAMLIN J., MILLER A. INTERFERENCE WITH IMMUNE HEMOLYSIS BY GLYCOPROTEIN ANTIGENS. J Exp Med. 1964 Nov 1;120:841–856. doi: 10.1084/jem.120.5.841. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Arquilla E. R., Bromer W. W., Mercola D. Immunology conformation and biological activity of insulin. Diabetes. 1969 Apr;18(4):193–205. doi: 10.2337/diab.18.4.193. [DOI] [PubMed] [Google Scholar]
  3. Atassi M. Z., Singhal R. P. Conformational studies on modified proteins and peptides. 3. Conformation of peptides obtained by cleavage of myoglobin at arginine peptide bonds. J Biol Chem. 1970 Oct 10;245(19):5122–5128. [PubMed] [Google Scholar]
  4. COWIE D. B., SPIEGELMAN S., ROBERTS R. B., DUERKSEN J. D. Ribosome-bound beta-galactosidase. Proc Natl Acad Sci U S A. 1961 Jan 15;47:114–122. doi: 10.1073/pnas.47.1.114. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cook R. A., Koshland D. E., Jr Specificity in the assembly of multisubunit proteins. Proc Natl Acad Sci U S A. 1969 Sep;64(1):247–254. doi: 10.1073/pnas.64.1.247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Fowler A. V., Zabin I. Beta-galactosidase: immunological studies of nonsense, missense and deletion mutants. J Mol Biol. 1968 Apr 14;33(1):35–47. doi: 10.1016/0022-2836(68)90279-9. [DOI] [PubMed] [Google Scholar]
  7. Fowler A. V., Zabin I. Co-linearity of beta-galactosidase with its gene by immunological detection of incomplete polypeptide chains. Science. 1966 Nov 25;154(3752):1027–1029. doi: 10.1126/science.154.3752.1027. [DOI] [PubMed] [Google Scholar]
  8. 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]
  9. Goldberg M. E. Tertiary structure of Escherichia coli beta-D-galactosidase. J Mol Biol. 1969 Dec 28;46(3):441–446. doi: 10.1016/0022-2836(69)90187-9. [DOI] [PubMed] [Google Scholar]
  10. HERZENBERG L. A. Studies on the induction of beta-galactosidase in a cryptic strain of Escherichia coli. Biochim Biophys Acta. 1959 Feb;31(2):525–538. doi: 10.1016/0006-3002(59)90029-0. [DOI] [PubMed] [Google Scholar]
  11. HORIUCHI T., TOMIZAWA J. I., NOVICK A. Isolation and properties of bacteria capable of high rates of beta-galactosidase synthesis. Biochim Biophys Acta. 1962 Jan 22;55:152–163. doi: 10.1016/0006-3002(62)90941-1. [DOI] [PubMed] [Google Scholar]
  12. KIHO Y., RICH A. INDUCED ENZYME FORMED ON BACTERIAL POLYRIBOSOMES. Proc Natl Acad Sci U S A. 1964 Jan;51:111–118. doi: 10.1073/pnas.51.1.111. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  14. Malkin L. I., Rich A. Partial resistance of nascent polypeptide chains to proteolytic digestion due to ribosomal shielding. J Mol Biol. 1967 Jun 14;26(2):329–346. doi: 10.1016/0022-2836(67)90301-4. [DOI] [PubMed] [Google Scholar]
  15. Morrison S. L., Zipser D. Polypeptide products of nonsense mutations. I. Termination fragments from nonsense mutations in the Z gene of the lac operon of Escherichia coli. J Mol Biol. 1970 Jun 14;50(2):359–371. doi: 10.1016/0022-2836(70)90198-1. [DOI] [PubMed] [Google Scholar]
  16. Taniuchi H., Anfinsen C. B. An experimental approach to the study of the folding of staphylococcal nuclease. J Biol Chem. 1969 Jul 25;244(14):3864–3875. [PubMed] [Google Scholar]
  17. ZIPSER D. STUDIES ON THE RIBOSOME-BOUND BETA-GALACTOSIDASE OF ESCHERICHIA COLI. J Mol Biol. 1963 Dec;7:739–751. doi: 10.1016/s0022-2836(63)80120-5. [DOI] [PubMed] [Google Scholar]
  18. Zomzely C. E., Roberts S., Peache S., Brown D. M. Cerebral protein synthesis. 3. Developmental alteratons in the stability of cerebral messenger ribonucleic acid-ribosome complexes. J Biol Chem. 1971 Apr 10;246(7):2097–2103. [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