The dimeric transmembrane domain of prolyl dipeptidase DPP-IV contributes to its quaternary structure and enzymatic activities.
Dipeptidyl peptidase IV (DPP-IV) is a drug target in the treatment of human type II diabetes. It is a type II membrane protein with a single transmembrane domain (TMD) anchoring the extracellular catalytic domain to the membrane. DPP-IV is active as a dimer, with two dimer interacting surfaces located extracellularly. In this study, we demonstrate that the TM of DPP-IV promotes DPP-IV dimerization and rescues monomeric DPP-IV mutants into partial dimers, which is specific and irreplaceable by TMs of other type II membrane proteins. By bioluminescence resonance energy transfer (BRET) and peptide electrophoresis, we found that the TM domain of DPP-IV is dimerized in mammalian cells and in vitro. The TM dimer interaction is very stable, based on our results with TM site-directed mutagenesis. None of the mutations, including the introduction of two prolines, resulted in their complete disruption to monomers. However, these TM proline mutations result in a significant reduction of DPP-IV enzymatic activity, comparable to what is found with mutations near the active site. A systematic analysis of TM structures deposited in the Protein Data Bank showed that prolines in the TM generally produce much bigger kinking angles than occur in nonproline-containing TMs. Thus, the proline-dependent reduction in enzyme activity may result from propagated conformational changes from the TM to the extracellular active site. Our results demonstrate that TM dimerization and conformation contribute significantly to the structure and activity of DPP-IV. Optimal enzymatic activity of DPP-IV requires an optimal interaction of all three dimer interfaces, including its TM.
Chung
Kuei-Min
KM
Division of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Zhu Nan town, Miaoli County, Taiwan, Republic of China.
Cheng
Jai-Hong
JH
Suen
Ching-Shu
CS
Huang
Chih-Hsiang
CH
Tsai
Cheng-Han
CH
Huang
Li-Hao
LH
Chen
Yi-Rong
YR
Wang
Andrew H-J
AH
Jiaang
Weir-Torn
WT
Hwang
Ming-Jing
MJ
Chen
Xin
X
D000595:Amino Acid Sequence; D000818:Animals; D018819:Dipeptidyl Peptidase 4 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment
D018819:Dipeptidyl Peptidase 4