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A G-quadruplex DNA-affinity Approach for Purification of Enzymatically Active G4 Resolvase1
Published on: March 18, 2017
Purification of recombinant poly(ADP-ribose) polymerases
Jean-Christophe Amé1, Thomas Kalisch, Françoise Dantzer
1Groupe Poly(ADP-ribosyl)ation et Intégrité du Génome, FRE3211 du CNRS, École Supérieure de Biotechnologie de, Strasbourg, France.
Methods in Molecular Biology (Clifton, N.J.)
|August 27, 2011
Summary
This study presents a rapid, three-step chromatographic protocol for purifying Poly(ADP-ribose) polymerases (PARPs). The enhanced method yields highly pure and active PARPs, including over 11 mg of mouse PARP-2 from 1 L of insect cell culture.
Area of Science:
- Biochemistry
- Molecular Biology
- Protein Purification
Background:
- Poly(ADP-ribose) polymerases (PARPs) are crucial enzymes involved in DNA repair and other cellular processes.
- Efficient purification of active PARPs is essential for biochemical and structural studies.
Purpose of the Study:
- To develop a fast, reproducible, and high-yield protocol for purifying PARPs.
- To obtain highly active PARP enzymes for further research.
Main Methods:
- A three-step chromatographic purification strategy was employed.
- Utilized Heparine Sepharose, 3-aminobenzamide Sepharose affinity chromatography, and Source 15S cation exchange chromatography.
- Purification was performed using an ÄKTA purifier system.
Main Results:
- The protocol successfully purified PARPs from overexpressing Sf9 insect cells and Escherichia coli.
- Achieved high purity and activity of PARP enzymes.
- Obtained over 11 mg of pure and active mouse PARP-2 per liter of Sf9 cell culture within 3 days.
Conclusions:
- The developed three-step chromatographic protocol offers a significant improvement for PARP purification.
- This method provides a reliable and efficient means to obtain substantial quantities of active PARPs.
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