Phage P4 origin-binding domain structure reveals a mechanism forregulation of DNA-binding activity by homo- and heterodimerization ofwinged helix proteins.
Mol Microbiol. 2002; 43: 855-67
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The origin-binding domain of the gpalpha protein of phage P4 (P4-OBD)mediates origin recognition and regulation of gpalpha activity by theprotein Cnr. We have determined the crystal structure of P4-OBD at 2.95 Aresolution. The structure of P4-OBD is that of a dimer with pseudo twofoldsymmetry. Each subunit has a winged helix topology with a unique structureamong initiator proteins. The only structural homologue of the P4-OBDsubunit is the DNA-binding domain of the eukaryotic transcriptionalactivator Rfx1. Based on this structural alignment, a model for originrecognition by the P4-OBD dimer is suggested. P4-OBD mutations thatinterfere with Cnr binding locate to the dimer interface, indicating thatCnr acts by disrupting the gpalpha dimer. P4-OBD dimerization is mediatedby helices alpha1 and alpha3 in both subunits, a mode of winged helixprotein dimerization that is reminiscent of that of the eukaryotictranscription factors E2F and DP. This, in turn, suggests that Cnr is alsoa winged helix protein, a possibility that is supported by previouslyunreported sequence homologies between Cnr and Rfx1 and homologymodelling. Hence, in a mechanism that appears to be conserved from phageto man, the DNA-binding activity of winged helix proteins can be regulatedby other winged helix proteins via the versatile use of the winged helixmotif as a homo- or heterodimerization scaffold.
The vaccinia virus D5 protein, which is required for DNA replication, is anucleic acid-independent nucleoside triphosphatase.
J Virol. 1995; 69: 5353-61
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The vaccinia virus D5 gene encodes a 90-kDa protein that is transientlyexpressed at early times after infection. Temperature-sensitive mutantswith lesions in the D5 gene exhibit a fast-stop DNA- phenotype and arealso impaired in homologous recombination. Here we report theoverexpression of the D5 protein within the context of a vaccinia virusinfection and its purification to apparent homogeneity. The purifiedprotein has an intrinsic nucleoside triphosphatase activity which isindependent of, and not stimulated by, any common nucleic acid cofactors.All eight common ribo- and deoxyribonucleoside triphosphates arehydrolyzed to the diphosphate form in the presence of a divalent cation.Implications for the role of D5 in viral DNA replication are addressed.
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