PMID:10212225
| Citation |
Mechanic, LE, Hall, MC and Matson, SW (1999) Escherichia coli DNA helicase II is active as a monomer. J. Biol. Chem. 274:12488-98 |
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| 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 (Kd = 3.4 microM), the protein is active in vivo and in vitro as a monomer. A mutant lacking the C-terminal 40 amino acids (UvrDDelta40C) failed to dimerize and yet was as active as the wild-type protein in ATP hydrolysis and helicase assays. In addition, the uvrDDelta40C 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. |
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| Keywords |
Adenosine Triphosphatases/chemistry; Adenosine Triphosphatases/genetics; Adenosine Triphosphatases/metabolism; Base Sequence; Chromatography, Gel; DNA Helicases; DNA Primers; DNA Repair; DNA, Single-Stranded/metabolism; Dimerization; Enzyme Activation; Escherichia coli/enzymology; Escherichia coli Proteins; Protein Binding |
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