Published in Journal of Molecular Biology
The Escherichia coli RecA protein has been a model for understanding homologous eukaryotic recombination proteins such as Rad51. The active form of both RecA and Rad51 appear to be helical filaments polymerized on DNA, in which an unusual helical structure is induced in the DNA. Surprisingly, the human meiosis-specific homolog of RecA, Dmc1, has th...
Published in Journal of Molecular Biology
We have investigated the biochemical properties of several Escherichia coli mutant recA proteins that display a null phenotype. These are the recA1, recA13 and recA56 proteins, each of which carries a single missense mutation. These proteins all share a common defect which is the inability to adopt the high affinity DNA binding state normally elici...
Published in Annual Review of Biophysics and Bioengineering
Published in The EMBO Journal
RecA protein features two distinct DNA-binding sites. During DNA strand exchange, the primary site binds to single-stranded DNA (ssDNA), forming the helical RecA nucleoprotein filament. The weaker secondary site binds double-stranded DNA (dsDNA) during the homology search process. Here we demonstrate that this site has a second important function. ...
Published in Nature Protocols
In this protocol, we describe a procedure to generate DNA dumbbells -single molecules of DNA with a microscopic bead attached at each end-and techniques for manipulating individual DNA dumbbells. We also detail the design and fabrication of a microfluidic device (flow cell) used in conjunction with dual optical trapping to manipulate DNA dumbbells...
Published in Cell
The breast cancer susceptibility protein, BRCA2, is essential for recombinational DNA repair. BRCA2 delivers RAD51 to double-stranded DNA (dsDNA) breaks through interaction with eight conserved, approximately 35 amino acid motifs, the BRC repeats. Here we show that the solitary BRC4 promotes assembly of RAD51 onto single-stranded DNA (ssDNA), but n...
Published in Journal of Molecular Biology
We have investigated quantitative molecular aspects of the interaction of recA protein with single-stranded DNA, by using a fluorescent modified-DNA referred to as etheno-M13 DNA. In addition, the effects of the nucleotide cofactors ATP and ADP, and the analogues ATP-gamma-S, AMP-P-C-P, and AMP-P-N-P on this interaction have been studied. It is sho...
Published in Nucleic Acids Research
DNA strand exchange, the central step of homologous recombination, is considered to occur approximately independently of DNA sequence content. However, certain prokaryotic and eukaryotic genomic loci display either an enhanced or reduced frequency of genetic exchange. Here we show that the Homo sapiens DNA strand exchange protein, HsRad51, shows a ...
Published in Nature
Published in Genes & Development
Homologous recombination in Escherichia coli is stimulated at DNA sequences known as chi sites. Stimulation requires the multifunctional RecBCD enzyme, which is both a helicase and a 3 —> 5 exonuclease. Upon recognition of a properly oriented chi site, the 3 —> 5 exonuclease activity is attenuated. Here we show that in addition to attenuation o...
