title: Molecular dissection of Mycobacterium tuberculosis integration host factor reveals novel insights into the mode of DNA binding and nucleoid compaction.
abstract: The annotated whole-genome sequence of Mycobacterium tuberculosis revealed that Rv1388 (Mtihf) is likely to encode for a putative 20-kDa integration host factor (mIHF). However, very little is known about the functional properties of mIHF or the organization of the mycobacterial nucleoid. Molecular modeling of the mIHF three-dimensional structure, based on the cocrystal structure of Streptomyces coelicolor IHF duplex DNA, a bona fide relative of mIHF, revealed the presence of Arg-170, Arg-171, and Arg-173, which might be involved in DNA binding, and a conserved proline (Pro-150) in the tight turn. The phenotypic sensitivity of Escherichia coli ΔihfA and ΔihfB strains to UV and methyl methanesulfonate could be complemented with the wild-type Mtihf but not its alleles bearing mutations in the DNA-binding residues. Protein-DNA interaction assays revealed that wild-type mIHF, but not its DNA-binding variants, binds with high affinity to fragments containing attB and attP sites and curved DNA. Strikingly, the functionally important amino acid residues of mIHF and the mechanism(s) underlying its binding to DNA, DNA bending, and site-specific recombination are fundamentally different from that of E. coli IHFαβ. Furthermore, we reveal novel insights into IHF-mediated DNA compaction depending on the placement of its preferred binding sites; mIHF promotes DNA compaction into nucleoid-like or higher order filamentous structures. We therefore propose that mIHF is a distinct member of a subfamily of proteins that serve as essential cofactors in site-specific recombination and nucleoid organization and that these findings represent a significant advance in our understanding of the role(s) of nucleoid-associated proteins.
authors.affiliation: From the Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Narayanaswamy
authors.initials: N
authors.lastname: Sharadamma
authors.affiliation: From the Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Yadumurthy
authors.initials: Y
authors.lastname: Harshavardhana
authors.affiliation: From the Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Apoorva
authors.initials: A
authors.lastname: Ravishankar
authors.affiliation: From the Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Praveen
authors.initials: P
authors.lastname: Anand
authors.affiliation: From the Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Nagasuma
authors.initials: N
authors.lastname: Chandra
authors.affiliation: From the Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India kmbc@biochem.iisc.ernet.in.
authors.forename: K
authors.initials: K
authors.lastname: Muniyappa
keywords: DNA Structure; Microbial Pathogenesis; Molecular Docking; Mycobacterium tuberculosis; Nucleic Acid Structure