Optimization of recombinant human poly(ADP-ribose) polymerase 3 production
https://doi.org/10.55959/MSU0137-0952-16-80-3S-12
Abstract
PARP3 (Poly(ADP-ribose) polymerase 3), like other PARP family members (PARP1 and PARP2), is an important factor in DNA repair. The specific functions and molecular mechanisms of action of this protein remain insufficiently studied. The development of a reliable protocol for obtaining high-purity and yield PARP3 is essential for comprehensive protein analysis, enzymatic activity, including studies of its binding to DNA, interactions with other protein factors, and structural investigations. In this study, we present a modified protocol for the expression of human PARP3 from Escherichia coli cells and its subsequent purification, which significantly increases protein yield compared to previously published methods.
About the Authors
E. A. Volkova
Department of Bioengineering, Faculty of Biology, Lomonosov Moscow State University
Russian Federation
Russian Federation
1–12 Leninskie Gory, Moscow, 119234
A. N. Korovina
Department of Bioengineering, Faculty of Biology, Lomonosov Moscow State University
Russian Federation
Russian Federation
1–12 Leninskie Gory, Moscow, 119234
N. V. Maluchenko
Department of Bioengineering, Faculty of Biology, Lomonosov Moscow State University
Russian Federation
Russian Federation
1–12 Leninskie Gory, Moscow, 119234
V. M. Studitsky
Department of Bioengineering, Faculty of Biology, Lomonosov Moscow State University; Fox Chase Cancer Center
Russian Federation
Russian Federation
1–12 Leninskie Gory, Moscow, 119234
333 Cottman Ave., Philadelphia, 19111, Pennsylvania
References
1. Augustin A., Spenlehauer C., Dumond H., Ménissier-de Murcia J., Piel M., Schmit A.C., Apiou F., Vonesch J.L., Kock M., Bornens M., De Murcia G. PARP-3 localizes preferentially to the daughter centriole and interferes with the G1/S cell cycle progression. J. Cell Sci. 2003;116(8):1551–1562.
2. Vyas S., Chesarone-Cataldo M., Todorova T., Huang Y.H., Chang P. A systematic analysis of the PARP protein family identifies new functions critical for cell physiology. Nat. Commun. 2013;4(1):2240.
3. Luo X., Kraus W.L. On PAR with PARP: cellular stress signaling through poly(ADP-ribose) and PARP-1. Genes Dev. 2012;26(5):417–432.
4. Langelier M.F., Riccio A.A., Pascal J.M. PARP-2 and PARP-3 are selectively activated by 5′ phosphorylated DNA breaks through an allosteric regulatory mechanism shared with PARP-1. Nucleic Acids Res. 2014;42(12):7762–7775.
5. Los M., Mozoluk M., Ferrari D., Stepczynska A., Stroh C., Renz A., Herceg Z., Wang Z.Q., Schulze-Osthoff K. Activation and caspase-mediated inhibition of PARP: A molecular switch between fibroblast necrosis and apoptosis in death receptor signaling. Mol. Biol. Cell. 2002;13(3):978–988.
6. Gupte R., Liu Z., Kraus W.L. PARPs and ADP-ribosylation: recent advances linking molecular functions to biological outcomes. Genes Dev. 2017;31(2):101–126.
7. Van Beek L., McClay É., Patel S., Schimpl M., Spagnolo L., Maia De Oliveira T. PARP Power: A structural perspective on PARP1, PARP2, and PARP3 in DNA damage repair and nucleosome remodelling. Int. J. Mol. Sci. 2021;22(10):5112.
8. Morales J., Li L., Fattah F.J., Dong Y., Bey E.A., Patel M., Gao J., Boothman D.A. Review of poly (ADP-ribose) polymerase (PARP) mechanisms of action and rationale for targeting in cancer and other diseases. Crit. Rev. Eukaryot. Gene Expr. 2014;24(1):15–28.
9. Rulten S.L., Fisher A.E.O., Robert I., Zuma M.C., Rouleau M., Ju L., Poirier G., Reina-San-Martin B., Caldecott K.W. PARP-3 and APLF function together to accelerate nonhomologous end-joining. Mol. Cell. 2011;41(1):33–45.
10. Langelier M.F., Steffen J.D., Riccio A.A., McCauley M., Pascal J.M. Purification of DNA damage-dependent PARPs from E. coli for structural and biochemical analysis. Poly(ADP-Ribose) Polymerase. Methods in Molecular Biology, vol. 1608. Ed. A.V. Tulin. N.Y.: Springer New York; 2017:431–444.
11. Amé J.C., Camuzeaux B., Dantzer F., Schreiber V. Purification of recombinant human PARP-3. Poly(ADP-Ribose) Polymerase. Methods in Molecular Biology, vol. 1608. Eds. A.V. Tulin. N.Y.: Springer New York; 2017:373–394.
12. Lowary P.T., Widom J. New DNA sequence rules for high affinity binding to histone octamer and sequence-directed nucleosome positioning. J. Mol. Biol. 1998;276(1):19–42.
13. Luger K., Rechsteiner T.J., Richmond T.J. Preparation of nucleosome core particle from recombinant histones. Chromatin. Methods in Enzymology, vol. 304. Eds. P.M. Wassarman and A.P. Wolffe. Academic Press; 1999:3–19.
14. Grundy G.J., Polo L.M., Zeng Z., Rulten S.L., Hoch N.C., Paomephan P., Xu Y., Sweet S.M., Thorne A.W., Oliver A.W., Matthews S.J., Pearl L.H., Caldecott K.W. PARP3 is a sensor of nicked nucleosomes and monoribosylates histone H2BGlu2. Nat. Commun. 2016;7(1):12404.
Review
For citations:
Volkova E.A., Korovina A.N., Maluchenko N.V., Studitsky V.M. Optimization of recombinant human poly(ADP-ribose) polymerase 3 production. Vestnik Moskovskogo universiteta. Seriya 16. Biologiya. 2025;80(3):81-86. (In Russ.) https://doi.org/10.55959/MSU0137-0952-16-80-3S-12
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