Purification and characterization of a new 120 kDa alkaline proteinase of Trypanosoma cruzi

Biochem Biophys Res Commun. 1992 Sep 30;187(3):1466-73. doi: 10.1016/0006-291x(92)90467-y.

Abstract

A new alkaline proteinase activity was identified in cell-free extracts of Trypanosoma cruzi epimastigotes on the basis of its ability to hydrolyze the fluorogenic substrate N-Z-Gly-Gly-Arg-AMC. The optimal activity was at pH 8.0. After a three step-chromatography procedure using two anionic columns (DEAE-Sepharose and Mono Q) and a chromatofocusing column (Mono P), the proteolytic activity was associated with a single 120 kDa protein and was called Tc 120 proteinase. The molecular mass of the proteinase was confirmed by direct visualization of the proteolytic activity using a fluorometric assay on SDS-PAGE. The Tc 120 proteinase which also cleaves N-Z-Arg-AMC, N-Z-Phe-Arg-AMC and N-glutaryl-Gly-Arg-AMC substrates, is a cysteine-type proteinase with an unusual low sensitivity to E-64.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Cysteine Endopeptidases / chemistry
  • Cysteine Endopeptidases / isolation & purification*
  • Cysteine Endopeptidases / metabolism
  • Hydrogen-Ion Concentration
  • Molecular Sequence Data
  • Molecular Weight
  • Peptides / metabolism
  • Protease Inhibitors / pharmacology
  • Protozoan Proteins / chemistry
  • Protozoan Proteins / isolation & purification
  • Protozoan Proteins / metabolism
  • Substrate Specificity
  • Trypanosoma cruzi / enzymology*

Substances

  • Peptides
  • Protease Inhibitors
  • Protozoan Proteins
  • Cysteine Endopeptidases
  • Tc 120 proteinase