Title

Specificity of DNA Binding and Dimerization of CspE and CspE Mutants from Escherichia Coli

Defense Date

4-19-2007

Graduation Date

Spring 1-1-2007

Availability

Campus Only

Submission Type

dissertation

Degree Name

PhD

Department

Biological Sciences

School

Bayer School of Natural and Environmental Sciences

Committee Chair

Nancy Trun

Committee Member

Joseph McCormick

Committee Member

Jana Patton-Vogt

Committee Member

Sue Wickner

Keywords

Cold-Shock Protein, CspE, Tryptophan Quenching, ssDNA-Binding, E. Coli, Dimerization

Abstract

Bacteria and cren-archeaota condense their chromosomes into compact nucleoids without the use of histone proteins or nucleosomes. To accomplish this, they use several different types of proteins, including topoisomerase, structural maintenance of chromosome (SMC)-like proteins, and small DNA binding and bending proteins. A small, DNA-binding protein involved in DNA compaction in E. coli K12 is CspE. In vivo, CspE has been shown to play a role in chromosome condensation and gene regulation. I have shown that CspE binds to ssDNA in vitro containing six continuous dT residues with high affinity, but will also bind to a lesser extent to other ssDNA templates. Unlike MukB, the major condensing protein, DNA binding by CspE is independent of ATP and divalent cations. The minimal length requirement for CspE binding to dT-stretches is eight nucleotides. CspE binding to dT-stretches in ssDNA is mainly resistant to high salt concentrations, implying that the interactions between CspE and ssDNA are non-ionic and are probably through hydrogen bonding and hydrophobic interactions. Several mutations in cspE have been shown to genetically separate the gene regulation phenotypes from the DNA architecture phenotypes, indicating a direct role for CspE in DNA condensation. In vitro biochemical characterization of these mutants has aided in the establishment of a model for chromosome condensation by CspE.

Format

PDF

Language

English

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