Studies of PARP-1 activation and inhibition using NMR spectroscopy

Poly(ADP-ribose)polymerase 1 (PARP-1) is a highly abundant multi-domain chromatin-associated enzyme found in all higher eukaryotic cell nuclei. It is the founding member of the PARP family of enzymes that modify themselves and other proteins by adding negatively charged poly(ADP-ribose) chains derived from NAD+. PARP-1 is a key sensor of DNA strand breaks and becomes automodified in response to DNA damage. PARP-1 activation occurs following recognition of single or double strand breaks by PARP-1’s N-terminal zinc finger domains. A multi-domain allosteric activation pathway leads to conformational changes in the HD subdomain of the catalytic domain that promote increased catalytic activity and lead to PARP-1 automodification and recruitment of DNA damage response proteins. Small molecule inhibitors of PARP-1 catalytic domain activity are also the first compounds to be discovered which target tumour cells with deficiencies in homologous recombination via a synthetically lethal approach. PARP-1 inhibitors cause cytotoxicity by preventing PARP-1 from being released from sites of DNA damage, but the mechanistic basis of the wide differences in effectiveness between different compounds has yet to be definitively established. Therefore, the aims of the work described in this thesis are to further elucidate the changes that occur in the HD subdomain during catalytic domain activation, and to help establish how binding of different PARP inhibitors to the catalytic domain in solution leads to the formation of trapped PARP-1 lesions using NMR spectroscopy.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2019

Author dc:creator

• Ogden, Tom

Advisor dc:contributor.advisor

• Neuhaus, David

• PARP-1
• PARP inhibitors
• PARP inhibition
• DNA damage response
• ADP-ribose
• ADP-ribosylation
• NMR spectroscopy
• Biomolecular NMR

Statement dc:rights

• Page 7, Figure 1.3. Reprinted from Molecular Cell, 58, Barkauskaite, E., Jankevicius, G., and Ahel, I, Structures and Mechanisms of Enzymes Employed in the Synthesis and Degradation of PARP-Dependent Protein ADP-Ribosylation, 935-946., Copyright (2015), with permission from Elsevier. Page 12, Figure 1.7. Reprinted from Molecular Cell, 58, Eustermann, S., Wu, W.-F., Langelier, M.-F., Yang, J.-C., Easton, L.E., Riccio, A.A., Pascal, J.M., and Neuhaus, D, Structural Basis of Detection and Signaling of DNA Single-Strand Breaks by Human PARP-1, 742-754., Copyright (2015), with permission from Elsevier. Page 13, Figure 1.8. Reprinted from Molecular Cell, 58, Dawicki-McKenna, J.M., Langelier, M.-F., DeNizio, J.E., Riccio, A.A., Cao, C.D., Karch, K.R., McCauley, M., Steffen, J.D., Black, B.E., and Pascal, J.M, PARP-1 Activation Requires Local Unfolding of an Autoinhibitory Domain, 755-768., Copyright (2015), with permission from Elsevier. Page 76, Figure 4.13. Reprinted from Molecular Cell, 58, Dawicki-McKenna, J.M., Langelier, M.-F., DeNizio, J.E., Riccio, A.A., Cao, C.D., Karch, K.R., McCauley, M., Steffen, J.D., Black, B.E., and Pascal, J.M, PARP-1 Activation Requires Local Unfolding of an Autoinhibitory Domain, 755-768., Copyright (2015), with permission from Elsevier. Appendix Page 4, Appendix Figure 1. From Hedgethorne, K., Eustermann, S., Yang, J.-C., Ogden, T.E.H., Neuhaus, D., and Bloomfield, G. (2017). Homeodomain-like DNA binding proteins control the haploid-to-diploid transition in Dictyostelium. Science Advances 3, e1602937.. Reprinted with permission from AAAS.

Licence

• https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/b30c7f25-9535-4d7b-81d5-1d3dc2d35610/download
• https://creativecommons.org/licenses/by/4.0/

Language dc:language

en

DOI dc:identifier.doi

https://doi.org/10.17863/CAM.47897

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/300822