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PMID: 270673 Published · ppublish English Journal Article

Reovirus mRNA can be covalently crosslinked via the 5' cap to proteins in initiation complexes.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 74 ·No. 10 ·1977-10-00 ·Pages 4288-92

Sonenberg N, Shatkin AJ

Abstract

Proteins that are located adjacent to the 5' end of mRNA in initiation complexes have been detected by chemical crosslinking. Reovirus mRNA containing radioactivity exclusively in the [3H]methyl-labeled "cap," m7G(5')ppp(5')-Gm, was oxidized with sodium periodate to convert the 2',3'-cis-diol of the 5'-terminal m7G to a reactive dialdehyde. Oxidized mRNA was incubated in cell-free protein-synthesizing systems derived from wheat germ or mammalian cells, and the resulting mRNA-ribosome initiation complexes were reduced with NaBH3CN. By this chemical procedure, putative Schiff bases between mRNA 5'termini and amino groups of neighboring proteins were stabilized by reduction, yielding covalently linked protein-RNA conjugates. Under conditions of ribosome binding, a limited number of polypeptides associated with the mRNA-ribosome complexes were crosslinked, suggesting that these proteins are positioned near and may interact with the 5' end of mRNA during initiation. This method should also be useful for studying the spatial relationships between molecules in other similar nucleoprotein complexes.

MeSH Terms
Animals In Vitro Techniques Oxidation-Reduction Peptide Initiation Factors/metabolism Peptides/metabolism Plant Extracts/metabolism Protein Binding RNA, Messenger/metabolism RNA, Viral/metabolism Rabbits Reoviridae/metabolism Reticulocytes/metabolism Ribosomes/metabolism
Chemicals
Peptide Initiation Factors Peptides Plant Extracts RNA, Messenger RNA, Viral
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Sonenberg N
Shatkin A J
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31 references, click to expand
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Article Info
Journal
Proceedings of the National Academy of Sciences of the United States of America
Abbr.
Proc Natl Acad Sci U S A
ISSN
0027-8424
DOI
10.1073/pnas.74.10.4288
Published
1977-10-00
Pages
4288-92
Language
English
Region
United States
NLM ID
7505876
PMCID
PMC431925
Subset
IM
External Links
PubMed source DOI: 10.1073/pnas.74.10.4288 PMC Full Text View journal details
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