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

Yohei Shiina,Hye-In J Lee,Takamasa Yoshizawa,Shinya Tsukiji,Koichi Takimoto,Ahmad Baihaqie,Zhili Zhang,Tomohiro Muto,Eulsoon Park

doi:10.1038/srep26290

Abstract

Mammalian DPP6 (DPPX) and DPP10 (DPPY) belong to a family of dipeptidyl peptidases, but lack enzyme activity. Instead, these proteins form complexes with voltage-gated K+ channels in Kv4 family to control their gating and other properties. Here, we find that the fly DPP10 ortholog acts as an ancillary subunit of Kv4 channels and digests peptides. Similarly to mammalian DPP10, the fly ortholog tightly binds to rat Kv4.3 protein. The association causes negative shifts in voltage dependence of channel activation and steady state inactivation. It also results in faster inactivation and recovery from inactivation. In addition to its channel regulatory role, fly DPP10 exhibits significant dipeptidyl peptidase activity with Gly-Pro-MCA (glycyl-L-proline 4-methylcoumaryl-7-amide) as a substrate. Heterologously expressed Flag-tagged fly DPP10 and human DPP4 show similar Km values towards this substrate. However, fly DPP10 exhibits approximately a 6-times-lower relative kcat value normalized with anti-Flag immunoreactivity than human DPP4. These results demonstrate that fly DPP10 is a dual functional protein, controlling Kv4 channel gating and removing bioactive peptides.

Highlights

DPP4-related proteins constitute a small group of dipeptidyl peptidases that are capable of hydrolyzing a prolyl bond two residues from the N-terminus
Our previous work demonstrated that this region of the two non-enzyme proteins mediate specific binding to Kv4 proteins[11]. These findings indicate that mammalian DPP6 and DPP10 act solely as auxiliary subunits of Kv4 channels, but not dipeptidyl peptidase
We searched for the presence of DPP6 and DPP10 orthologs in various species using HomoloGene (NCBI)

Summary

Introduction

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 DPP91–3 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 disorders[4,5,6]. 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 family[9,10,11,12]. Our previous work demonstrated that this region of the two non-enzyme proteins mediate specific binding to Kv4 proteins[11] These findings indicate that mammalian DPP6 and DPP10 act solely as auxiliary subunits of Kv4 channels, but not dipeptidyl peptidase. We show that the fly DPP10 ortholog possesses a dual function as channel-ancillary subunit and peptidase

Methods

Results

Conclusion