Purification and biochemical characterization of dipeptidyl peptidase-II (DPP7) homologue from germinated Vigna radiata seeds

• This is the first detailed study of DPP-II from plants.
• Plant DPP-II is a serine protease which preferably cleaves Lys-Ala-4mβNA at pH 7.5.
• It is a oligomeric enzyme with molecular weight of approximately 97.3 kDa.
• These findings open interest in the study of involvement of this enzyme in plant development.
• Plant DPP-II may be involved in activation of other proteases and bioactive peptide generation.

Dipeptidyl peptidases (DPPs) are potent exopeptidases, which possess central role in proteolysis. As compared to other members of DPP family, proline containing dipeptide hydrolysing activity of DPP-II (Dipeptidyl peptidase II) is unique as it hydrolyses imino group and plays a key role in protein metabolism. In present study, DPP-II was purified from germinated moong bean seeds using acid and ammonium sulphate precipitation followed by successive chromatographies on gel filtration (pH 7.4) and cation exchanger (pH 5.9). Native PAGE and in-situ gel assay confirmed the apparent homogeneity. Purified plant DPP-II is an oligomeric enzyme with molecular weight of 97.3 kDa. Highest DPP-II activity was observed at pH 7.5 and 37 °C, with stability in the range of neutral to alkaline pH. Substrate specificity showed consequent activity for proline containing dipeptide followed by Lys-Ala and other hydrophobic dipeptides, but none of the studied endopeptidase and monopeptidase substrate was hydrolysed. Catalytic characterization with modifier studies revealed the involvement of Ser and His residues in its catalytic mechanism. Its dipeptidyl peptidase activity for proline containing dipeptide supported its role in the bioactive peptide generation and food industry. Functional studies of DPP-II revealed the significant involvement of this glycoproteinous enzyme in protein mobilization during germination. Further studies on industrial applications exploring physiological role are in progress.

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