Purification and Characterization of DNA Helicase BstH2 from Bacillus Stearothermophilus.

ABSTRACT

In the purification of DNA helicase BstH1 we have partially purified the second DNA helicase BstH2 from Bacillus Stearothermophilus through Polymin P precipitation, ammonia sulfate precipitation and chromatographic steps with DEAE-cellulose, phosphocellulose, Blue-Sepharose, FPLC Superose 12, Mono Q and second Mono Q. The ATPase activity of BstH2 depends on Mg(2+) and is differentially stimulated by different types of nucleic acids. BstH2 has a maximal ATPase activity at 55 degrees. The ATPase activity is greatly inhibited by E. coli SSB or higher ionic strength. The DNA helicase activity of BstH2 depends on ATP and Mg(2+). BstH2 can unwind partial duplex DNA as well as blunt-ended duplex DNA. E. coli SSB stimulates the unwinding reaction catalyzed by BstH2.