Rapid monitoring of autolysis process of proteases by capillary electrophoresis

Biotechnol Lett. 2003 Oct;25(20):1763-7. doi: 10.1023/a:1026040012947.

Abstract

A protease, MCP-01, produced by a deep-sea psychrotrophic strain of Pseudoaltermonas sp. SM9913 was purified and its autolysis reaction at 20 degrees C-50 degrees C was monitored by capillary electrophoresis. Capillary electrophoresis provides a rapid assay because the degree and state of autolysis of protease MCP-01 could be observed within 6 min. The autolysis rate increased as the temperature rose in the tested range. After 30 min incubation at 30 degrees C, 77% of MCP-01 autolyzed into peptides. However, its activity for the hydrolysis of casein was reduced by only 4%. The rate of loss of activity of MCP-01 was thus slower than that of autolysis of MCP-01 at 30 degrees C. Similar results were obtained when MCP-01 was incubated at 20 degrees C, 40 degrees C and 50 degrees C. Large peptides produced by autolysis of MCP-01 therefore still have catalytic activity. When these large peptides autolyzed further into smaller peptides, the enzyme conformation that retained its catalytic activity was destroyed and activity was lost.

Publication types

  • Evaluation Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Autolysis*
  • Electrophoresis, Capillary / methods*
  • Endopeptidases / analysis*
  • Endopeptidases / chemistry*
  • Enzyme Activation
  • Protein Conformation
  • Protein Denaturation
  • Quality Control
  • Temperature*

Substances

  • Endopeptidases
  • protease MCP-01, Pseudoaltermonas