RecA protein promoted DNA strand exchange.

D A Soltis,I R Lehman

doi:10.1016/s0021-9258(18)32374-3

Abstract

recA protein and circular single-stranded DNA form a stable complex in the presence of single-stranded DNA binding protein (SSB), in which one recA protein monomer is bound per two nucleotides of DNA. These complexes are kinetically significant intermediates in the exchange of strands between the single-stranded DNA and an homologous linear duplex. After completion of strand exchange, the recA protein remains tightly associated with the circular duplex product of the reaction and the SSB is bound to the displaced linear single strand. Upon addition of ADP, the recA protein-duplex DNA complex dissociates. RecA protein also interacts with single-stranded DNA in the absence of SSB; however, the amount of recA protein bound is substantially reduced. These findings provide direct physical evidence for the participation of SSB in the formation of the recA protein-single-stranded DNA complexes inferred earlier from kinetic analysis. Moreover, they confirm the ability of recA protein to equilibrate between bound and free forms in the absence of SSB.

Highlights

RecA protein and circular single-strandedDNA form Electron microscopic studies have demonstrated that recA a stablecomplexinthepresenceofsingle-stranded protein will form large, filamentous aggregatesin the presence
Protein-duplexDNA complex dissociatesR. ecA protein interacts with single-strandeDdNA in the absence of SSB; theamount of recA protein boundis substantially reduced. These findings provide direct physical evidence for the participation of SSB in the formation of therecAprotein-single-stranded DNA complexes inferred earliefrom kinetic analysisM. oreover, they confirm the abilityof recA protein to equilibrate between bound anfdree forms in the absenceof SSB
When an aliquot was predicted from earlier studies which showed that after from the reaction was analyzedby agarose gel electrophoresis, the completion of strand exchange, recA protein was unable to react with added SS DNA when the ATP/ADP ratio was maintained at sufficiently high levels [8]

Summary

Introduction

RecA protein and circular single-strandedDNA form Electron microscopic studies have demonstrated that recA a stablecomplexinthepresenceofsingle-stranded protein will form large, filamentous aggregatesin the presence. RecA monomer is bound per two nucleotides of DNA. These protein forms highly ordered structures with singlecomplexes are kinetically significant intermediates in strandedand double-stranded DNA [9,10,11,12].Possibly, such the exchange of strands between the single-stranded structures are related to the complexesinvolved in strand. SS DNA complexesand analyzed tightly associated with the circular duplexproduct of the substrates, intermediates, and products of a DNA strand thereactionandthe SSB is bound tothedisplaced linear single strand. Of SSB; theamount of recA protein boundis substantially reduced. These findings provide direct physical evidence for the participation of SSB in the formation of therecAprotein-single-stranded DNA complexes inferred earliefrom kinetic analysisM. McMacken, Johns Hopkins University, using a modification of a published procedure

Methods

Results

Conclusion