Fly DPP10 acts as a channel ancillary subunit and possesses peptidase activity

DPP4-related proteins constitute a small group of dipeptidyl peptidases that are capable of hydrolyzing a prolyl bond two residues from the N-terminus. The group includes DPP4, FAP (fibroblast activation protein), DPP8 and DPP91,2,3. Since these enzymes potentially degrade important hormones, cytokines and other proteins, they have attracted much interest as pharmacological targets for treatment of metabolic, immune and other disorders4,5,6. In contrast to these enzymes, the two structurally-related members DPP6 (DPPX) and DPP10 (DPPY) lack peptidase activity7,8. Indeed, the corresponding polypeptides contain aspartic acid and glycine, respectively, at the position of the catalytic serine in DPP4-related enzymes. Instead, these two proteins tightly bind to pore-forming subunits of voltage-gated K+ channels in Kv4 family9,10,11,12. The channel complexes containing DPP6 or DPP10 display various gating properties that resemble fast-inactivating A-type K+ currents in native neurons13,14. Thus, mammalian DPP6 and DPP10 act as auxiliary subunits for neuronal Kv4 channels. Crystal structures of mammalian DPP4, DPP6 and DPP10 reveal the common structural organization in their extracellular regions with a unique eight-blade β propeller and α/β-hydrolase fold15,16,17. The structural studies also indicated that the lack of enzymatic activity in mammalian DPP6 and DPP10 was not simply the result of the amino acid substitutions at the catalytic serine16,17. The positions of several other residues and functional groups in the catalytic triad and substrate-recognizing portion show notable differences between the two non-enzyme proteins and DPP4. Thus, mammalian DPP6 and DPP10 might have lost various structural features required for peptidase activity during evolution. Moreover, the transmembrane and its surrounding region of DPP6 and DPP10 polypeptides show significant sequence divergence from the corresponding portion of DPP4 protein. Our previous work demonstrated that this region of the two non-enzyme proteins mediate specific binding to Kv4 proteins11. These findings indicate that mammalian DPP6 and DPP10 act solely as auxiliary subunits of Kv4 channels, but not dipeptidyl peptidase. Channel ancillary subunits might arise from proteins with other functional roles during evolution. In some cases, these proteins may retain the original or related function while gaining a new channel-regulatory role. Furthermore, the original or related functions may control gating and other properties of the associated channel. For instance, Kvβ proteins are functional oxidoreductases that act as Kv1-channel ancillary subunits18,19. Mutations in the catalytic or NADPH-binding sites, as well as the cofactor binding itself, influence the ability of Kvβ subunit to alter channel inactivation20,21,22. Moreover, oxidation of NADPH on Kvβ has been shown to influence inactivation of the associated channel23. Thus, Kvβ subunit may act as a redox sensor to control excitability. We wondered if any channel-interacting peptidase homologs might retain enzymatic activity, potentially acting as a link between the enzymatic function to excitability. Inspection of primary structures of dipeptidyl peptidases and their homologs in the GenBank detected potential dual functional molecules in non-mammalian species. In this paper, we show that the fly DPP10 ortholog possesses a dual function as channel-ancillary subunit and peptidase.