Abstract
Protein poly(ADP-ribosyl)ation (PARylation) regulates a number of important cellular processes. Poly(ADP-ribose) glycohydrolase (PARG) is the primary enzyme responsible for hydrolyzing the poly(ADP-ribose) (PAR) polymer in vivo. Here we report crystal structures of the mouse PARG (mPARG) catalytic domain, its complexes with ADP-ribose (ADPr) and a PARG inhibitor ADP-HPD, as well as four PARG catalytic residues mutants. With these structures and biochemical analysis of 20 mPARG mutants, we provide a structural basis for understanding how the PAR polymer is recognized and hydrolyzed by mPARG. The structures and activity complementation experiment also suggest how the N-terminal flexible peptide preceding the PARG catalytic domain may regulate the enzymatic activity of PARG. This study contributes to our understanding of PARG catalytic and regulatory mechanisms as well as the rational design of PARG inhibitors.
Highlights
Protein function and localization inside the cell are usually regulated by post-translational modifications (PTMs)
PARylation is catalyzed by a family of poly(ADP-ribose) polymerases (PARPs), which modify the target protein side chains by transferring the ADP-ribose (ADPr) moiety from nicotinamide adenine dinucleotide (NAD+)
Implications in poly(ADP-ribose) glycohydrolase (PARG) catalytic mechanism Based on the bacterial, protozoan, rat and human PARG
Summary
Protein function and localization inside the cell are usually regulated by post-translational modifications (PTMs). Poly(ADPribosyl)ation (PARylation) is a reversible PTM that is involved in various cellular processes, including DNA repair, chromatin structure dynamics, gene transcription, poly(ADP-ribose) (PAR) dependent cell death (pathanatos) and PARylation dependent ubiquitination [1,2,3,4,5]. Reversal of PARylation is predominantly carried out by poly(ADP-ribose) glycohydrolase (PARG) in nucleus and cytosol, whereas ADPribosylhydrolase 3 (ARH3) may play a role in mitochondrial PAR degradation [6]. PARPs and PARG are present in all eukaryotic cell types except yeast [7]. The 111 kDa full length PARG (hPARG111) localizes in the nucleus. Both 99 kDa hPARG99 and 102 kDa hPARG102 isoforms localize in the cytoplasm. While the N-terminal region is absent in some PARG splicing forms and predicted to be disordered (Fig. S1) [9], the conserved C-terminal 60 kD catalytic domain is fully active [10,11]
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