Enhancement of recA protein-promoted DNA strand exchange activity by volume-occupying agents.

摘要: To investigate the in vivo effects of macromolecular crowding we examined effect inert macromolecules such as polyvinyl alcohol and polyethylene glycol on vitro activity recA protein. The addition either these volume-occupying agents enables protein to promote homologous pairing exchange DNA strands at an otherwise nonpermissive magnesium ion concentration. In presence macromolecules, both rate association with single-stranded (ssDNA) steady-state affinity for ssDNA are increased. Consequently, ability compete ssDNA-binding (SSB protein) is enhanced, inhibitory SSB formation protein-ssDNA presynaptic complexes eliminated. Because bind ssDNA-containing secondary structures also enhanced volume-occupied solution, joint molecule not greatly reduced when omitted from reaction. Thus, increased interactions contribute complex formation. addition, must affect another property protein, i.e. self-association, which required synapsis strand exchange. Our examination helps reconcile requirement elevated concentrations protein-promoted recombination reactions vitro, a presumably low concentration vivo.