Purification and characterization of a protease from the visceral mass of Mytella charruana and its evaluation to obtain antimicrobial peptides

Food Chem. 2018 Apr 15:245:1169-1175. doi: 10.1016/j.foodchem.2017.11.044. Epub 2017 Nov 14.

Abstract

This work describes purification of a protease from the visceral mass of the mussel Mytella charruana as well as evaluation of its ability to hydrolyze milk casein to generate antimicrobial peptides. The enzyme showed pI of 4.1 and a single polypeptide band of 83.1 kDa after SDS-PAGE. Sequence similarities with tropomyosin and myosin from mollusks were detected. The protease showed a trypsin-like activity with optimal temperature of 40 °C and stability in a wide pH range (3.0-9.0). Km was 4.28 ± 0.34 mM of the synthetic substrate N-benzoyl-dl-arginyl-ρ-nitroanilide, whereas Vmax was 0.056 ± 0.001 nmol min-1. The enzyme hydrolyzed casein, and the hydrolysate inhibited the growth of Escherichia coli, Micrococcus luteus, Bacillus subtilis, and Klebsiella pneumoniae at a minimal inhibitory concentration of 5.0 µg mL-1. In conclusion, the visceral mass of M. charruana contains a trypsin-like protease that can generate peptides from casein that have a bacteriostatic effect.

Keywords: Antibacterial activity; Bioactive peptides; Bivalve mollusk; Casein; Protease.

MeSH terms

  • Animals
  • Anti-Bacterial Agents / chemistry
  • Anti-Bacterial Agents / pharmacology*
  • Antifungal Agents / chemistry
  • Antifungal Agents / pharmacology*
  • Bivalvia / enzymology*
  • Caseins / chemistry
  • Caseins / metabolism
  • Drug Evaluation, Preclinical / methods
  • Electrophoresis, Polyacrylamide Gel
  • Enzyme Stability
  • Hydrogen-Ion Concentration
  • Hydrolysis
  • Microbial Sensitivity Tests
  • Peptides / chemistry
  • Peptides / metabolism
  • Peptides / pharmacology*
  • Serine Endopeptidases / isolation & purification
  • Serine Endopeptidases / metabolism*
  • Temperature
  • Viscera / enzymology

Substances

  • Anti-Bacterial Agents
  • Antifungal Agents
  • Caseins
  • Peptides
  • trypsin-like serine protease
  • Serine Endopeptidases