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A G-quadruplex DNA-affinity Approach for Purification of Enzymatically Active G4 Resolvase1
Kapitel
- 00:05Titel
- 00:54Lyse Bacterial Pellets Expressing Human Recombinant G4 Resolvase1 (rG4R1)
- 03:06Prepare G4-magnetic Beads
- 04:07Bind Histidine-tagged rG4R1 to Cobalt Beads
- 05:47Elute rG4R1 from Cobalt Beads
- 07:07Bind rG4R1 to G4-bound SPB and Elute in an ATP-dependent Manner
- 08:55Results: Analysis and Quantification of Catalytically Active, Purified rG4R1
- 10:06Conclusion
G4 Resolvase1 binds to G-quadruplex (G4) structures with the tightest reported affinity for a G4-binding protein and represents the majority of the G4-DNA unwinding activity in HeLa cells. We describe a novel protocol that harnesses the affinity and ATP-dependent unwinding activity of G4-Resolvase1 to specifically purify catalytically active recombinant G4R1.
