Human cathepsin F: expression in baculovirus system, characterization and inhibition by protein inhibitors

Biol Chem. 2004 Jun;385(6):505-9. doi: 10.1515/BC.2004.059.

Abstract

Recombinant full-length human procathepsin F, produced in the baculovirus expression system, was partially processed during the purification procedure to a form lacking the N-terminal cystatin-like domain and activated with pepsin. Active cathepsin F efficiently hydrolyzed Z-FR-MCA (kcat/Km=106 mM(-1) s(-1)) and Bz-FVR-MCA (kcat/Km=8 mM(-1) s(-1)), whereas hydrolysis of Z-RR-MCA was very slow (kcat/Km<0.2 mM(-1) s(-1)). Cathepsin F was rapidly and tightly inhibited by cystatin C, chicken cystatin and equistatin with Ki values in the subnanomolar range (0.03-0.47 nM), whereas L-kininogen was a less strong inhibitor of the enzyme (Ki=4.7 nM). Stefin A inhibited cathepsin F slowly (kass=1.6 x 10(5) M(-1) s(-1)) and with a lower affinity (Ki=25 nM). These data suggest that cathepsin F differs from other related endopeptidases by considerably weaker inhibition by stefins.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Baculoviridae / genetics*
  • Baculoviridae / metabolism
  • Cathepsin F
  • Cathepsins / antagonists & inhibitors*
  • Cathepsins / chemistry*
  • Cathepsins / genetics
  • Cell Line
  • Enzyme Activation
  • Gene Expression
  • Gene Transfer Techniques
  • Humans
  • Hydrolysis
  • Kinetics
  • Protease Inhibitors / pharmacology*
  • Recombinant Proteins / antagonists & inhibitors
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Spodoptera

Substances

  • Protease Inhibitors
  • Recombinant Proteins
  • Cathepsins
  • CTSF protein, human
  • Cathepsin F