Abstract
Tankyrases are poly(ADP-ribose)polymerases (PARPs) which recognize their substrates via their ankyrin repeat cluster (ARC) domains. The human tankyrases (TNKS/TNKS2) contain five ARCs in their extensive N-terminal region; of these, four bind peptides present within tankyrase interactors and substrates. These short, linear segments, known as tankyrase-binding motifs (TBMs), contain some highly conserved features: an arginine at position 1, which occupies a predominantly acidic binding site, and a glycine at position 6 that is sandwiched between two aromatic side chains on the surface of the ARC domain. Tankyrases are involved in a multitude of biological functions, amongst them Wnt/β-catenin signaling, the maintenance of telomeres, glucose metabolism, spindle formation, the DNA damage response and Hippo signaling. As many of these are relevant to human disease, tankyrase is an important target candidate for drug development. With the emergence of non-catalytic (scaffolding) functions of tankyrase, it seems attractive to interfere with ARC function rather than the enzymatic activity of tankyrase. To study the mechanism of ARC-dependent recruitment of tankyrase binders and enable protein-observed NMR screening methods, we have as the first step obtained a full backbone and partial side chain assignment of TNKS2 ARC4. The assignment highlights some of the unusual structural features of the ARC domain.
Highlights
The tankyrases (TNKS/ARTD5, TNKS2/ARTD6) are poly(ADP-ribose)polymerases (PARPs) and as such catalyse the processive modification of protein substrates with poly(ADP-ribose) (PAR) chains, thereby consuming their co-substrate NAD+ (Haikarainen et al 2014)
The ankyrin repeat cluster (ARC) are followed by a polymerizing sterile alpha motif (SAM) domain (De Rycker and Price 2004; Mariotti et al 2016; Riccio et al 2016) that precedes the PARP domain (Lehtiö et al 2008)
Additional roles of tankyrase include the regulation of mitotic spindle formation (Chang et al 2005, 2009), Hippo signaling (Wang et al 2015; Troilo et al 2016; Jia et al 2017) and emerging functions in the DNA damage response (Nagy et al 2016), cell migration (Lupo et al 2016), and Notch signaling (Bhardwaj et al 2017), to name a few
Summary
The tankyrases (TNKS/ARTD5, TNKS2/ARTD6) are poly(ADP-ribose)polymerases (PARPs) and as such catalyse the processive modification of protein substrates with poly(ADP-ribose) (PAR) chains, thereby consuming their co-substrate NAD+ (Haikarainen et al 2014). A second key binding determinant is a channel lined by two aromatic amino acids which accommodates an essential glycine residue at TBM position 6, forming an ‘aromatic glycine sandwich’ (Guettler et al 2011) Their strict requirement suggests that the arginine and glycine residues represent critical docking points in substrate recruitment. There is evidence for tankyrase targets devoid of detectable TBM sequences (Li et al 2017; Bhardwaj et al 2017) These binders may either indirectly interact with tankyrase or be recruited through direct binding by alternative, hitherto unknown binding mechanisms. Targeting tankyrase’s accessory domains provides an attractive alternative means to inhibit tankyrase function This approach would block both tankyrase-dependent scaffolding and substrate PARylation. We report the assignment of ARC4 of human TNKS2, which will enable a more elaborate characterisation of substrate recruitment by tankyrase ARCs and the development of substrate binding antagonists
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