Characterization and Crystal Structure of Prolyl Endopeptidase from Abalone (Haliotis discus hannai)
Wan-Yu Li, Yue Li, Yu-Lei Chen, Jian-Jian Hu, Hylemariam Mihiretie Mengist, Guang-Ming Liu, Tengchuan Jin, Min-Jie Cao
Aimed to study the characteristics of prolyl endopeptidase (PEP, EC 22.214.171.124) and its possible role in the degradation of collagen, we cloned the full-length cDNA sequence of PEP from abalone (Haliotis discus hannai) (Hdh-PEP). Recombinant Hdh-PEP (rHdh-PEP) was expressed in vitro, its enzymatic properties were detected, and its secondary structure was analyzed by Circular Dichroism (CD). We for the first time determined the 1.5 Å crystal structure of rHdh-PEP. The decomposition effect of rHdh-PEP on collagen peptides was analyzed. Our data revealed that the molecular weight of rHdh-PEP is 85 kDa, consisting of a catalytic domain and a β-propeller domain. The optimal pH and temperature of rHdh-PEP were pH 6.0 and 20˚C, respectively. Using small collagen peptides as substrates, HPLC-ESI-MS analysis confirmed that rHdh-PEP specifically cleaved at the carboxyl side of proline residues, suggesting its role in the degradation of collagen peptides during autolysis.
Fluorescence, Enzyme activity, Kinetics, Biochemistry