Linear dichroism study of RecA-DNA complexes. Structural evidence and binding stoichiometries

Abstract

In the presence of RecA single-stranded DNA (ssDNA) is found to exhibit flow linear dichroism (LD). In the absence of the cofactor ATP gamma S, the LD is positive with a maximum at about 280 nm, whereas in the presence of the cofactor ATP gamma S there is still a positive long-wavelength band, but a negative LD contribution centered at 260 nm indicates an orientation of the DNA bases preferentially perpendicular to the fiber axis. For the complex between ssDNA and RecA without ATP gamma S, essentially all LD derives from the protein (tryptophane) subunits indicating a structure in which the tryptophanes are preferentially parallel to the fiber axis of the complex while the DNA bases remain essentially unoriented. The magnitude of the LD increases with the RecA/DNA ratio to a point corresponding to approximately three nucleotides per RecA and decreases thereafter with excess of DNA. This indicates that there are two modes of binding with different stoichiometries.