title: Biochemical Characterization of a Mycobacteriophage Derived DnaB Ortholog Reveals New Insight into the Evolutionary Origin of DnaB Helicases.
abstract: The bacterial replicative helicases known as DnaB are considered to be members of the RecA superfamily. All members of this superfamily, including DnaB, have a conserved C- terminal domain, known as the RecA core. We unearthed a series of mycobacteriophage encoded proteins in which the RecA core domain alone was present. These proteins were phylogenetically related to each other and formed a distinct clade within the RecA superfamily. A mycobacteriophage encoded protein, Wildcat Gp80 that roots deep in the DnaB family, was found to possess a core domain having significant sequence homology (Expect value < 10-5) with members of this novel cluster. This indicated that Wildcat Gp80, and by extrapolation, other members of the DnaB helicase family, may have evolved from a single domain RecA core polypeptide belonging to this novel group. Biochemical investigations confirmed that Wildcat Gp80 was a helicase. Surprisingly, our investigations also revealed that a thioredoxin tagged truncated version of the protein in which the N-terminal sequences were removed was fully capable of supporting helicase activity, although its ATP dependence properties were different. DnaB helicase activity is thus, primarily a function of the RecA core although additional N-terminal sequences may be necessary for fine tuning its activity and stability. Based on sequence comparison and biochemical studies we propose that DnaB helicases may have evolved from single domain RecA core proteins having helicase activities of their own, through the incorporation of additional N-terminal sequences.
authors.affiliation: Department of Microbiology, Bose Institute, P1/12 C.I.T. Scheme VIIM, Kolkata 700054, West Bengal, India.
authors.forename: Priyanka
authors.initials: P
authors.lastname: Bhowmik
authors.affiliation: Department of Microbiology, Bose Institute, P1/12 C.I.T. Scheme VIIM, Kolkata 700054, West Bengal, India.
authors.forename: Sujoy K
authors.initials: SK
authors.lastname: Das Gupta
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D003001:Cloning, Molecular; D004277:DNA, Single-Stranded / Q000378:metabolism; D053488:DnaB Helicases / Q000737:chemistry / Q000145:classification / Q000235:genetics* / Q000378:metabolism; D019143:Evolution, Molecular; D006868:Hydrolysis; D009160:Mycobacteriophages / Q000201:enzymology* / Q000235:genetics; D009838:Oligodeoxyribonucleotides / Q000378:metabolism; D010802:Phylogeny; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011938:Rec A Recombinases / Q000145:classification; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D013879:Thioredoxins; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D004277:DNA, Single-Stranded; D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; D013879:Thioredoxins; D000255:Adenosine Triphosphate; D011938:Rec A Recombinases; D000251:Adenosine Triphosphatases; D053488:DnaB Helicases