University of Illinois Urbana-Champaign

Biochemical and genetic analysis of nucleic acid-protein interactions

Wang, Siqun

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Description

Title
Biochemical and genetic analysis of nucleic acid-protein interactions
Author(s)
Wang, Siqun
Issue Date
1995
Doctoral Committee Chair(s)
Gardner, Jeffrey F.
Department of Study
Microbiology
Discipline
Microbiology
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
  • Biology, Molecular
  • Biology, Genetics
  • Biology, Microbiology
Language
eng
Abstract
  • The integration host factor (IHF) of Escherichia coli is a small, sequence-specific DNA-binding protein. The specific and nonspecific DNA binding constants were determined by gel mobility shift assays. The binding constant of IHF for the H$\sp\prime$ site in $\lambda$ attP is 6.8 $\times$ 10$\sp8$ M$\sp{-1}$ and the nonspecific binding constant is 5.8 $\times$ 10$\sp5$ M$\sp{-1}.$ Therefore, the selectivity of IHF binding is about a thousand-fold higher for a specific site over random sequences. To study the molecular determinants specifying IHF binding, a series of 41 oligonucleotides were synthesized containing adenine analogues that modified the surfaces of the major and minor grooves of DNA. The binding constants determined for these analogue-containing H$\sp\prime$ sites revealed that the specific interaction of IHF with its H$\sp\prime$ site involves interactions with both minor and major grooves of the DNA.
  • The coat protein of coliphage R17 is a small, sequence-specific RNA-binding protein. The ribophage system was used to characterize the specific interactions between the coat protein and its RNA-binding site. Studies with the RNA-binding site suggest that the translational repression conferred by this hairpin depends on its interactions with both the R17 coat protein and the ribosome. In addition, a total of eight R17 coat protein mutants were isolated with altered, expanded RNA-binding activities in vivo. They contain substitutions at the N-terminus and in the middle of the coat protein. Molecular characterizations of these coat protein mutants suggest that there are two classes of mutants. Possible mechanisms of the in vivo suppression of phage variants by these mutant coat proteins are discussed.
Type of Resource
text
Permalink
http://hdl.handle.net/2142/21092
Copyright and License Information
Copyright 1995 Wang, Siqun

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Graduate Dissertations and Theses at Illinois PRIMARY
Graduate Theses and Dissertations at Illinois
Dissertations and Theses - Microbiology