Specific recognition of DNA by integration host factor. Glutamic acid 44 of the β-subunit specifies the discrimination of a T:A from an A:T base pair without directly contacting the DNA

Erik K. Read, Richard I. Gumport, Jeffrey F. Gardner

Research output: Contribution to journalArticlepeer-review

Abstract

Integration host factor (IHF) is a protein that binds to the H' site of bacteriophage λ with sequence specificity. Genetic experiments implicated amino acid residue Glu44 of the β-subunit of IHF in discrimination against substitution of A for T at position 44 of the TTR submotif of the binding site (Lee, E. C., Hales, L. M., Gumport, R. I., Gardner, J. F. (1992) EMBO J., 11, 305-313). We have extended this observation by generating all possible single-base substitutions at positions 43, 44, and 45 of the H' site. IHF failed to bind these H' site substitution mutants in vivo. The K(d)(app) value for each H' site substitution, except for H'45A mutant, was reduced >2000-fold relative to the wild-type site. Substitution of amino acid β-Glu44 with alanine prevented IHF from discriminating against the H'44A variant but not the other H' site substitution mutants. Further analysis with other substitutions at position β44 demonstrated that both oxygens of the wild-type glutamic acid are necessary for discrimination of AT at position 44. Because the β-Glu44 residue does not contact the DNA, this residue probably enforces binding specificity indirectly through interaction with amino acids that themselves contact the DNA.

Original languageEnglish (US)
Pages (from-to)33759-33764
Number of pages6
JournalJournal of Biological Chemistry
Volume275
Issue number43
DOIs
StatePublished - Oct 27 2000

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Fingerprint

Dive into the research topics of 'Specific recognition of DNA by integration host factor. Glutamic acid 44 of the β-subunit specifies the discrimination of a T:A from an A:T base pair without directly contacting the DNA'. Together they form a unique fingerprint.

Cite this