title: Mycobacterium tuberculosis DinG is a structure-specific helicase that unwinds G4 DNA: implications for targeting G4 DNA as a novel therapeutic approach.
abstract: The significance of G-quadruplexes and the helicases that resolve G4 structures in prokaryotes is poorly understood. The Mycobacterium tuberculosis genome is GC-rich and contains >10,000 sequences that have the potential to form G4 structures. In Escherichia coli, RecQ helicase unwinds G4 structures. However, RecQ is absent in M. tuberculosis, and the helicase that participates in G4 resolution in M. tuberculosis is obscure. Here, we show that M. tuberculosis DinG (MtDinG) exhibits high affinity for ssDNA and ssDNA translocation with a 5' → 3' polarity. Interestingly, MtDinG unwinds overhangs, flap structures, and forked duplexes but fails to unwind linear duplex DNA. Our data with DNase I footprinting provide mechanistic insights and suggest that MtDinG is a 5' → 3' polarity helicase. Notably, in contrast to E. coli DinG, MtDinG catalyzes unwinding of replication fork and Holliday junction structures. Strikingly, we find that MtDinG resolves intermolecular G4 structures. These data suggest that MtDinG is a multifunctional structure-specific helicase that unwinds model structures of DNA replication, repair, and recombination as well as G4 structures. We finally demonstrate that promoter sequences of M. tuberculosis PE_PGRS2, mce1R, and moeB1 genes contain G4 structures, implying that G4 structures may regulate gene expression in M. tuberculosis. We discuss these data and implicate targeting G4 structures and DinG helicase in M. tuberculosis could be a novel therapeutic strategy for culminating the infection with this pathogen.
authors: From the Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
authors: Roshan Singh
authors: RS
authors: Thakur
authors: From the Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
authors: Ambika
authors: A
authors: Desingu
authors: From the Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
authors: Shivakumar
authors: S
authors: Basavaraju
authors: From the Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
authors: Shreelakshmi
authors: S
authors: Subramanya
authors: From the Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
authors: Desirazu N
authors: DN
authors: Rao
authors: From the Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India nganesh@biochem.iisc.ernet.in.
authors: Ganesh
authors: G
authors: Nagaraju
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D055162:Biocatalysis; D002942:Circular Dichroism; D018983:DNA Footprinting / Q000379:methods; D004265:DNA Helicases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D045566:DNA, Cruciform / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D054856:G-Quadruplexes; D006801:Humans; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D014376:Tuberculosis / Q000188:drug therapy / Q000382:microbiology
chemical_list: D001426:Bacterial Proteins; D045566:DNA, Cruciform; D004277:DNA, Single-Stranded; D004791:Enzyme Inhibitors; D003850:Deoxyribonuclease I; D004265:DNA Helicases; C000594764:DinG protein, Mycobacterium tuberculosis
keywords: DNA Damage Response; DNA Helicase; DNA Recombination; DNA Repair; DNA-Protein Interaction