Escherichia coli DNA helicase II is active as a monomer

Abstract

Helicases are thought to function as oligomers (generally dimers or hexamers). Here we demonstrate that although Escherichia coli DNA helicase II (UvrD) is capable of dimerization as evidenced by a positive interaction in the yeast two-hybrid system, gel filtration chromatography, and equilibrium sedimentation ultracentrifugation (K(d) = 3.4 μM), the protein is active in vivo and in vitro as a monomer. A mutant lacking the C-terminal 40 amino acids (UvrDΔ40C) failed to dimerize and yet was as active as the wild-type protein in ATP hydrolysis and helicase assays. In addition, the uvrDΔ40C allele fully complemented the loss of helicase II in both methyl-directed mismatch repair and excision repair of pyrimidine dimers. Biochemical inhibition experiments using wild-type UvrD and inactive UvrD point mutants provided further evidence for a functional monomer. This investigation provides the first direct demonstration of an active monomeric helicase, and a model for DNA unwinding by a monomer is presented.