title: Fluorescent sensors of PARP-1 structural dynamics and allosteric regulation in response to DNA damage.
abstract: Poly(ADP-ribose) (PAR) is a posttranslational modification predominantly synthesized by PAR polymerase-1 (PARP-1) in genome maintenance. PARP-1 detects DNA damage, and damage detection is coupled to a massive increase PAR production, primarily attached to PARP-1 (automodification). Automodified PARP-1 then recruits repair factors to DNA damage sites. PARP-1 automodification eventually leads to release from DNA damage thus turning off catalytic activity, although the effects of PAR on PARP-1 structure are poorly understood. The multiple domains of PARP-1 are organized upon detecting DNA damage, creating a network of domain contacts that imposes a major conformational transition in the catalytic domain that increases PAR production. Presented here are two novel fluorescent sensors that monitor the global and local structural transitions of PARP-1 that are associated with DNA damage detection and catalytic activation. These sensors display real-time monitoring of PARP-1 structural transitions upon DNA damage detection, and their reversal upon PARP-1 automodification. The fluorescent sensors are further used to investigate intramolecular and intermolecular PARP-1 activation, followed by the observation that intramolecular activation of PARP-1 is the predominant response to DNA strand breaks in cells. These results provide a unique perspective on the interplay between PARP-1 DNA damage recognition, allosteric regulation, and catalytic activity.
authors.affiliation: Department of Biochemistry & Molecular Biology, Sidney Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, PA 19107, USA.
authors.forename: Jamin D
authors.initials: JD
authors.lastname: Steffen
authors.affiliation: Department of Biochemistry & Molecular Biology, Sidney Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, PA 19107, USA.
authors.forename: Michael M
authors.initials: MM
authors.lastname: McCauley
authors.affiliation: Department of Biochemistry & Molecular Biology, Sidney Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, PA 19107, USA john.pascal@umontreal.ca.|Department of Biochemistry & Molecular Medicine, Université de Montréal, Montréal, QC H3T 1J4, Canada.
authors.forename: John M
authors.initials: JM
authors.lastname: Pascal
mesh_terms: D000494:Allosteric Regulation; D000818:Animals; D015374:Biosensing Techniques; D002384:Catalysis; D002460:Cell Line; D004249:DNA Damage; D004789:Enzyme Activation; D004795:Enzyme Stability; D031541:Fluorescence Resonance Energy Transfer; D051379:Mice; D008958:Models, Molecular; D009243:NAD / Q000737:chemistry / Q000378:metabolism; D000071137:Poly (ADP-Ribose) Polymerase-1 / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D054730:Protein Interaction Domains and Motifs; D013329:Structure-Activity Relationship
chemical_list: D009243:NAD; D000071137:Poly (ADP-Ribose) Polymerase-1