title: DnaJ (Hsp40 protein) binding to folded substrate impacts KplE1 prophage excision efficiency.
abstract: Temperate phages mediate gene transfer and can modify the properties of their host organisms through the acquisition of novel genes, a process called lysogeny. The KplE1 prophage is one of the 10 prophage regions in Escherichia coli K12 MG1655. KplE1 is defective for lysis but fully competent for site-specific recombination. The TorI recombination directionality factor is strictly required for prophage excision from the host genome. We have previously shown that DnaJ promotes KplE1 excision by increasing the affinity of TorI for its site-specific recombination DNA target. Here, we provide evidence of a direct association between TorI and DnaJ using in vitro cross-linking assays and limited proteolysis experiments that show that this interaction allows both proteins to be transiently protected from trypsin digestion. Interestingly, NMR titration experiments showed that binding of DnaJ involves specific regions of the TorI structure. These regions, mainly composed of α-helices, are located on a surface opposite the DNA-binding site. Taken together, we propose that DnaJ, without the aid of DnaK/GrpE, is capable of increasing the efficiency of KplE1 excision by causing a conformational stabilization that allows TorI to adopt a more favorable conformation for binding to its specific DNA target.
authors.affiliation: Laboratoire de Chimie Bactérienne, CNRS UMR7283, Institut de Microbiologie de la Méditerranée, Aix-Marseille University, 31 Chemin Joseph Aiguier, 13402 Marseille Cedex 20, France.
authors.forename: Tania M
authors.initials: TM
authors.lastname: Puvirajesinghe
authors.forename: Latifa
authors.initials: L
authors.lastname: Elantak
authors.forename: Sabrina
authors.initials: S
authors.lastname: Lignon
authors.forename: Nathalie
authors.initials: N
authors.lastname: Franche
authors.forename: Marianne
authors.initials: M
authors.lastname: Ilbert
authors.forename: Mireille
authors.initials: M
authors.lastname: Ansaldi
mesh_terms: D001665:Binding Sites; D002942:Circular Dichroism; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000378:metabolism*; D050956:HSP40 Heat-Shock Proteins / Q000378:metabolism*; D008242:Lysogeny; D013058:Mass Spectrometry / Q000379:methods; D008954:Models, Biological; D018832:Molecular Chaperones / Q000378:metabolism; D011418:Prophase; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011995:Recombination, Genetic; D013379:Substrate Specificity; D014357:Trypsin / Q000737:chemistry / Q000494:pharmacology; D014775:Virus Activation
chemical_list: D003432:Cross-Linking Reagents; C494635:DnaJ protein, E coli; D029968:Escherichia coli Proteins; D050956:HSP40 Heat-Shock Proteins; D018832:Molecular Chaperones; C488589:TorI protein, E coli; D014357:Trypsin