The ATPase, RNA unwinding, and RNA binding activities of recombinant p68 RNA helicase.
p68 RNA helicase, a nuclear RNA helicase, was identified 2 decades ago. The protein plays very important roles in cell development and organ maturation. However, the biological functions and enzymology of p68 RNA helicase are not well characterized. We report the expression and purification of recombinant p68 RNA helicase in a bacterial system. The recombinant p68 is an ATP-dependent RNA helicase. ATPase assays demonstrated that double-stranded RNA (dsRNA) is much more effective than single-stranded RNA in stimulating ATP hydrolysis by the recombinant protein. Consistently, RNA-binding assays showed that p68 RNA helicase binds single-stranded RNA weakly in an ATP-dependent manner. On the other hand, the recombinant protein has very high affinity for dsRNA. Binding of the protein to dsRNA is ATP-independent. The data indicate that p68 may directly target dsRNA as its natural substrate. Interestingly, the recombinant p68 RNA helicase unwinds dsRNA in both 3' --> 5' and 5' --> 3' directions. This is the second example of a Asp-Glu-Ala-Asp (DEAD) box RNA helicase that unwinds RNA duplexes in a bi-directional manner.
Huang
Youliang
Y
Program in Cell and Molecular Biosciences, Department of Animal and Dairy Sciences, Auburn University, 210 Upchurch Hall, Auburn, Alabama 36849, USA.
Liu
Zhi-Ren
ZR
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D001483:Base Sequence; D053487:DEAD-box RNA Helicases; D017931:DNA Primers; D004789:Enzyme Activation; D006868:Hydrolysis; D011485:Protein Binding; D011494:Protein Kinases / Q000378:metabolism*; D012313:RNA / Q000737:chemistry / Q000378:metabolism*; D020365:RNA Helicases / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism
D017931:DNA Primers; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D012313:RNA; D011494:Protein Kinases; D000251:Adenosine Triphosphatases; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases