DNA-BINDING AND HELICASE DOMAINS OF THE ESCHERICHIA-COLI RECOMBINATION PROTEIN RECG

The Escherichia coli RecG protein is a unique junction-specific helica se involved in DNA repair and recombination. The C-terminus of RecG co ntains motifs conserved throughout a wide range of DNA and RNA helicas es and it is thought that this C-terminal half of RecG contains the he licase active site. However, the regions of RecG which confer junction DNA specificity are unknown. To begin to assign structure-function re lationships within RecG, a series of N- and C-terminal deletions have been engineered into the protein, together with an N-terminal histidin e tag fusion peptide for purification purposes. Junction DNA binding, unwinding and ATP hydrolysis were disrupted by mutagenesis of the N-te rminus, In contrast, C-terminal deletions moderately reduced junction DNA binding but almost abolished unwinding. These data suggest that th e C-terminus does contain the helicase active site whereas the N-termi nus confers junction DNA specificity.