Further characterization and partial amino acid sequence of a cysteine proteinase from Trypanosoma cruzi.
A cysteine proteinase from epimastigotes of Trypanosoma cruzi, Tul 2 stock, has been purified to homogeneity from cell-free extracts obtained by freezing and thawing, by a procedure involving ammonium sulfate fractionation, DEAE-Sephacel chromatography, and gel filtration on Sephadex G-200; when necessary, further purification was attained by fast protein liquid chromatography on Mono Q and Superose 6 columns. The purified enzyme was strongly inhibited by leupeptin, antipain and chymostatin (I50 values of 0.25, 0.75 and 1 microM, respectively), little inhibited by elastatinal, and unaffected by pepstatin A. The enzyme is a glycoprotein, as shown by binding to ConA-Sepharose and elution with alpha-methyl-D-mannopyranoside and alpha-methyl-D-glucopyranoside. Partial amino acid sequences were obtained from the N-terminal end (32 amino acids) of the carbamidomethylated enzyme, and from a tryptic peptide (14 amino acids) of the pyridylethylated enzyme. Both regions show considerable homology with papain and some cathepsins, such as cathepsin L, thus showing that the enzyme belongs to the cysteine proteinase family.
Cazzulo
J J
JJ
Instituto de Investigaciones Bioquímicas Fundación Campomar, Buenos Aires, Argentina.
Couso
R
R
Raimondi
A
A
Wernstedt
C
C
Hellman
U
U
D000595:Amino Acid Sequence; D000818:Animals; D056668:Cathepsin L; D002403:Cathepsins; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D003546:Cysteine Endopeptidases / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases; D006023:Glycoproteins / Q000302:isolation & purification; D008969:Molecular Sequence Data; D010206:Papain; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D014349:Trypanosoma cruzi / Q000201:enzymology*
D006023:Glycoproteins; D011500:Protein Synthesis Inhibitors; D002403:Cathepsins; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; D056668:Cathepsin L; D010206:Papain