Purification, characterization and the use of recombinant prolyl oligopeptidase from Myxococcus xanthus for gluten hydrolysis

- Myxococcus xanthus prolyl oligopeptidase expressed in E.coli was purified 60.3 fold.
- Enzyme was characterized for its biochemical, catalytic properties and its regulation by using certain metal ions and chemicals.
- The potential use of the recombinant prolyl oligopeptidase was tested with wheat gluten hydrolysis.

Prolyl oligopeptidase (POP, EC 3.4.21.26) is a cytosolic serine protease that hydrolyses proline containing small peptides. The members of prolyl oligopeptidase family play important roles in many physiological processes such as neurodegenerative diseases, maturation and degradation of peptide hormones. Thus the enzyme has been purified and characterized from various sources to elucidate the potential use as therapeutics. In this study recombinant Myxococcus xanthus prolyl oligopeptidase expressed in E. coli was purified 60.3 fold, using metal-chelate affinity and gel permeation chromatography. The recombinant enzyme had a monomeric molecular weight of 70 kDa. Isoelectric point of the enzyme was found to be approximately 6.3 by two-dimensional polyacrylamide gel electrophoresis. The optimum pH and temperature was estimated as 7.5 and 37 °C, respectively. The purified enzyme was stable in a pH range of 6.0-8.5 and thermally stable up to 37 °C. The Km and Vmax values were 0.2 mM and 3.42 μmol/min/mg. The proteolytic activity was inhibited by active-site inhibitors of serine protease, Z-Pro-Prolinal, PMSF, and metal ions, Cd2+, and Hg2+. Furthermore, the hydrolysis efficiency of the recombinant prolyl oligopeptidase was investigated with wheat gluten.