Specific recognition nucleotides and their DNA context determine the affinity of E2 protein for 17 binding sites in the BPV-1 genome

Li, R., Knight, J., Bream, G., Stenlund, A., Botchan, M. (April 1989) Specific recognition nucleotides and their DNA context determine the affinity of E2 protein for 17 binding sites in the BPV-1 genome. Genes Dev, 3 (4). pp. 510-26. ISSN 0890-9369 (Print)0890-9369 (Linking)

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URL: http://www.ncbi.nlm.nih.gov/pubmed/2542129
DOI: 10.1101/gad.3.4.510

Abstract

The DNA context of nucleotides that a protein recognizes can influence the strength of the protein-DNA interaction. Moreover, in prokaryotes, understanding the quantitative differences in binding affinities that result in part from the DNA context is often important in describing regulatory mechanisms. Nevertheless, these issues have not been a major focus yet for the investigation of protein-DNA interactions in eukaryotes. In this study, we explored the binding specificity and the range of affinities that the BPV-1 E2 transcriptional activator has for DNA. Because E2 binding sites are positioned near several different BPV-1 promoters, such quantitative information may be important to understand transcriptional regulatory mechanisms in BPV-1. Gel retardation assays and DNA footprinting were used to quantitate the affinities of the E2 binding sites in the viral genome. In the process, five sites were discovered, which, on the basis of sequence, had not been predicted previously to interact with the E2 protein. Equilibrium and kinetic studies show that the range of E2 affinities of the 17 sites varied over 300-fold. The sequence elements responsible for E2 recognition of DNA were determined by missing contact analysis of several sites and a point mutation analysis of one site. The results presented show that the affinity of an E2 binding site is to a large extent determined by the availability of specific contacts, but the data also strongly suggest that DNA structure plays an important role.

Item Type: Paper
Uncontrolled Keywords: Animals Base Sequence Binding Sites Bovine papillomavirus 1/ genetics Cattle Computer Graphics DNA Probes DNA, Viral/ genetics/metabolism DNA-Binding Proteins/genetics/ metabolism Densitometry Deoxyribonuclease I/diagnostic use Gene Expression Regulation Molecular Sequence Data Mutation Nucleotides/ genetics Papillomaviridae/ genetics Viral Proteins/genetics/ metabolism
Subjects: bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification
bioinformatics > genomics and proteomics > genetics & nucleic acid processing
bioinformatics > genomics and proteomics
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein types > DNA binding protein
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein types
organism description > virus
CSHL Authors:
Communities: CSHL labs > Stenlund lab
Depositing User: Matt Covey
Date: April 1989
Date Deposited: 12 Mar 2013 18:48
Last Modified: 03 Nov 2017 21:02
Related URLs:
URI: http://repository.cshl.edu/id/eprint/27784

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