BACKGROUND Nucleotide excision repair is a versatile DNA repair reaction that removes bulky adducts generated by environmental mutagens such as the UV spectrum of sunlight or chemical carcinogens. Current multistep models of this excision repair pathway accommodate its broad substrate repertoire but fail to explain the stringent selectivity toward damaged nucleotides among excess native DNA. To...

We recently demonstrated that the RecBCD enzyme is a bipolar DNA helicase that employs two single-stranded DNA motors of opposite polarity to drive translocation and unwinding of duplex DNA. We hypothesized that this organization may explain the exceptionally high rate and processivity of DNA unwinding catalyzed by the RecBCD enzyme. Using a stopped-flow dye displacement assay for unwinding act...

Genetic evidence indicates that Saccharomyces cerevisiae Sgs1, Top3, and Rmi1 resolve topologically linked intermediates arising from DNA replication and recombination. Using purified proteins, we show that Sgs1, Top3, Rmi1, and replication protein A (RPA) coordinate catenation and decatenation of dsDNA through sequential passage of single strands of DNA, establishing a unique pathway for dsDNA...

In the preceding articles we have described the isolation and some of the properties of two calf thymus proteins which bind selectively to single-stranded DNA and which appear analogous to previously isolated prokaryotic DNA-unwinding proteins. In the present work we demonstrate two further points of analogy. First, both the calf UP1 and the high salt eluting proteins form protein-rich complexe...

In Arabidopsis thaliana, nuclear multisubunit RNA polymerase IV (Pol IV) and RNA-DEPENDENT RNA POLYMERASE 2 (RDR2) are required for the biogenesis of 24-nucleotide small interfering RNAs (siRNAs) that direct DNA methylation and transcriptional silencing at target loci transcribed by nuclear multisubunit RNA polymerase V (Pol V). Pol IV and RDR2 physically associate and RDR2's polymerase activit...

A DNA helicase, called chloroplast DNA (ctDNA) helicase II, was purified to apparent homogeneity from pea (Pisum sativum). The enzyme contained intrinsic, single-stranded, DNA-dependent ATPase activity and an apparent molecular mass of 78 kD on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The DNA helicase was markedly stimulated by DNA substrates with fork-like replication structu...

Alternative DNA structures that deviate from B-form double-stranded DNA such as G-quadruplex (G4) DNA can be formed by G-rich sequences that are widely distributed throughout the human genome. We have previously shown that Pif1p not only unfolds G4, but also unwinds the downstream duplex DNA in a G4-stimulated manner. In the present study, we further characterized the G4-stimulated duplex DNA u...

Bacteriophage T5 has a 120 kb double-stranded linear DNA genome encoding most of the genes required for its own replication. This lytic bacteriophage has a burst size of ∼500 new phage particles per infected cell, demonstrating that it is able to turn each infected bacterium into a highly efficient DNA manufacturing machine. To begin to understand DNA replication in this prodigious bacteriophag...

Following DNA strand exchange, recA protein remains associated with the heteroduplex DNA product of the reaction. This association exists without significant changes (as measured by nuclease protection, topological state of the heteroduplex DNA, and rates of ATP hydrolysis) for at least 30 min after strand exchange is complete. The heteroduplex DNA is unwound as a result of this association to ...