Structural features of adenovirus 2 virus-associated RNA required for binding to the protein kinase DAI

Clarke, P. A., Pe'ery, T., Ma, Y., Mathews, M. B. (October 1994) Structural features of adenovirus 2 virus-associated RNA required for binding to the protein kinase DAI. Nucleic Acids Res, 22 (21). pp. 4364-74. ISSN 0305-1048 (Print)

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URL: http://www.ncbi.nlm.nih.gov/pubmed/7971266
DOI: 10.1093/nar/22.21.4364

Abstract

The double-stranded RNA activated protein kinase DAI contains an RNA binding domain consisting of two copies of a double-stranded RNA binding motif. We have investigated the role of RNA structure in the interaction between DAI and the structured single-stranded RNA, adenovirus VA RNAI, which inhibits DAI activation. Mutations in the apical stem, terminal stem, and central domain of the RNA were tested to assess the contribution of these elements to DAI binding in vitro. The data demonstrate that over half a turn of intact apical stem is required for the interaction and that there is a correlation between the binding of apical stem mutants and their ability to function both in vivo and in vitro. There was also evidence of preference for GC-rich sequence in the proximal region of the apical stem. In the central domain the correlation between binding and function of mutant RNAs was poor, suggesting that at least some of this region plays no direct role in binding to DAI, despite its functional importance. Exceptionally, central domain mutations that encroached on the phylogenetically conserved stem 4 of VA RNA disrupted binding, and complementary mutations in this sequence partially restored binding. Measurement of the binding of wild-type VA RNAI to DAI and p20, a truncated form of the protein containing the RNA binding domains alone, under various ionic conditions imply that the major interactions are electrostatic and occur via the protein's RNA binding domain. However, differences between full-length DAI and p20 in their binding to mutants in the conserved stem suggest that regions outside the RNA binding domain also participate in the binding. The additional interactions are likely to be non-ionic, and may be important for preventing DAI activation during virus infection.

Item Type: Paper
Uncontrolled Keywords: Adenoviruses, Human/ genetics Base Sequence Binding Sites Comparative Study Electrochemistry Enzyme Activation Escherichia coli Gene Deletion Humans Molecular Sequence Data Mutagenesis Mutation Nucleic Acid Conformation Protein-Serine-Threonine Kinases/ metabolism RNA, Viral/ chemistry/genetics/metabolism Recombinant Proteins/metabolism Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. Structure-Activity Relationship eIF-2 Kinase
Subjects: bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > RNA expression
organism description > virus > adenovirus
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein types > enzymes > kinase
CSHL Authors:
Communities: CSHL labs
Depositing User: Matt Covey
Date: 25 October 1994
Date Deposited: 25 Aug 2015 19:32
Last Modified: 08 Nov 2017 16:00
PMCID: PMC308468
Related URLs:
URI: http://repository.cshl.edu/id/eprint/31430

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