The SUMO1-E67 Interacting Loop Peptide Is an Allosteric Inhibitor of the Dipeptidyl Peptidases 8 and 9

Abstract

The intracellular peptidases dipeptidyl peptidase (DPP) 8 and DPP9 are involved in multiple cellular pathways including antigen maturation, cellular homeostasis, energy metabolism, and cell viability. Previously we showed that the small ubiquitin-like protein modifier SUMO1 interacts with an armlike structure in DPP9, leading to allosteric activation of the peptidase. Here we demonstrate that the E67-interacting loop (EIL) peptide, which corresponds to the interaction surface of SUMO1 with DPP9, acts as a noncompetitive inhibitor of DPP9. Moreover, by analyzing the sensitivity of DPP9 arm mutants to the EIL peptide, we mapped specific residues in the arm that are important for inhibition by the EIL, suggesting that the peptide acts as an allosteric inhibitor of DPP9. By modifying the EIL peptide, we constructed peptide variants with more than a 1,000-fold selectivity toward DPP8 (147 nm) and DPP9 (170 nm) over DPPIV (200 μm). Furthermore, application of these peptides to cells leads to a clear inhibition of cellular prolyl peptidase activity. Importantly, in line with previous publications, inhibition of DPP9 with these novel allosteric peptide inhibitors leads to an increase in EGF-mediated phosphorylation of Akt. This work highlights the potential use of peptides that mimic interaction surfaces for modulating enzyme activity.Background: SUMO1 binds to an arm motif in the prolyl-peptidase DPP9, leading to allosteric activation of the peptidase.Results: A SUMO1 peptide covering the DPP9 interaction surface inhibits DPP9 activity. Inhibition is dependent on residues in the DPP9 arm motif.Conclusion: The SUMO1 peptide and its variants are allosteric DPP9 inhibitors.Significance: This work highlights the potential use of peptides mimicking interaction surfaces for modulating enzyme activity. The intracellular peptidases dipeptidyl peptidase (DPP) 8 and DPP9 are involved in multiple cellular pathways including antigen maturation, cellular homeostasis, energy metabolism, and cell viability. Previously we showed that the small ubiquitin-like protein modifier SUMO1 interacts with an armlike structure in DPP9, leading to allosteric activation of the peptidase. Here we demonstrate that the E67-interacting loop (EIL) peptide, which corresponds to the interaction surface of SUMO1 with DPP9, acts as a noncompetitive inhibitor of DPP9. Moreover, by analyzing the sensitivity of DPP9 arm mutants to the EIL peptide, we mapped specific residues in the arm that are important for inhibition by the EIL, suggesting that the peptide acts as an allosteric inhibitor of DPP9. By modifying the EIL peptide, we constructed peptide variants with more than a 1,000-fold selectivity toward DPP8 (147 nm) and DPP9 (170 nm) over DPPIV (200 μm). Furthermore, application of these peptides to cells leads to a clear inhibition of cellular prolyl peptidase activity. Importantly, in line with previous publications, inhibition of DPP9 with these novel allosteric peptide inhibitors leads to an increase in EGF-mediated phosphorylation of Akt. This work highlights the potential use of peptides that mimic interaction surfaces for modulating enzyme activity. Background: SUMO1 binds to an arm motif in the prolyl-peptidase DPP9, leading to allosteric activation of the peptidase. Results: A SUMO1 peptide covering the DPP9 interaction surface inhibits DPP9 activity. Inhibition is dependent on residues in the DPP9 arm motif. Conclusion: The SUMO1 peptide and its variants are allosteric DPP9 inhibitors. Significance: This work highlights the potential use of peptides mimicking interaction surfaces for modulating enzyme activity.