Linked Life Data

16225979

Source:http://linkedlifedata.com/resource/pubmed/id/16225979

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
1
pubmed:dateCreated
2005-11-25
pubmed:abstractText
The Escherichia coli dnaA73, dnaA721, and dnaA71 alleles, which encode A213D, R432L, T435K substitutions, respectively, were originally isolated as extragenic suppressors of a temperature-sensitive dnaX mutant. As the A213D substitution resides in a domain that functions in ATP binding and the R432L and T435K substitutions affect residues that recognize the DnaA box motif, they might be expected to reduce ATP and specific DNA binding, respectively. Therefore, a major objective was to quantify the biochemical defects of the mutant DnaAs to understand how the altered proteins suppress the temperature-sensitive phenotype of a dnaX mutant. A second purpose was to address the paradox that mutant proteins with substitutions of amino acids essential for recognition of the DnaA box motifs within the E. coli replication origin (oriC) may well be inactive in initiation, yet chromosomal dnaA mutants expressing DnaA proteins with the R432L and T435K substitutions are viable at temperatures from 30 to 39 degrees C. We show biochemically that mutant DnaAs carrying R432L and T435K substitutions fail to bind to the DnaA box sequence. The A213D mutant is sevenfold reduced in its affinity for ATP compared to wild-type DnaA, and its affinity for the DnaA box sequence is also reduced. However, the reduced activity of the A213D mutant in oriC plasmid replication appears to arise from a defect in DnaA oligomerization. Although the T435K mutant fails to bind to the DnaA box sequence, other results suggest that DnaA oligomerization stabilizes the binding of the mutant DnaA to oriC to support its partial activity in initiation in vitro. These results support a model that suppression of dnaX occurs by reducing the frequency of initiation to a manageable level for the mutant DnaX so that viability is maintained.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jan
pubmed:issn
0300-9084
pubmed:author
pubmed:issnType
Print
pubmed:volume
88
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
1-10
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
2006
pubmed:articleTitle
Escherichia coli DnaA protein: specific biochemical defects of mutant DnaAs reduce initiation frequency to suppress a temperature-sensitive dnaX mutation.
pubmed:affiliation
Section of Molecular Genetics and Microbiology, Institute for Cellular and Molecular Biology, University of Texas, Austin, TX 78712, USA. jrw@mail.utexas.edu
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't, Research Support, N.I.H., Extramural