Multiple protease activities in Giardia intestinalis trophozoites.
Azocasein and a wide range of chromogenic and fluorogenic peptidyl substrates, representing 21 different peptide sequences, were degraded by the intracellular proteases present in lysates of trophozoites of Giardia intestinalis (syn lamblia) Portland 1 strain. Total protease activity differed considerably with the substrate with the p-nitroanilide derivatives Bz-pro-phe-arg-pNA and Bz-phe-val-arg-pNA being most rapidly hydrolysed (specific activities 4.2 +/- 0.4 and 1.1 +/- 0.6 U/mg protein, respectively). Activities were increased (16-72%) by addition of 1 mM dithiothreitol and were maximal, for the substrates monitored, in the range pH 5.5-7.0. These data and the inhibitor susceptibilities of the trophozoite proteases confirmed that the activity was predominantly due to cysteine proteases, although the presence of some serine protease, aspartic protease and aminopeptidase activity in the lysates was also indicated. The multiplicity of the protease activities was confirmed by gelatin-polyacrylamide electrophoretic analysis. Eighteen proteolytic activities with Mr values in the range 30,000 to > 211,000 were detected. These gelatinase activities were enhanced by dithiothreitol, were maximal at or close to pH 6, and were inhibited by cysteine proteinase and some serine proteinase inhibitors. Four of the proteases present in the gels exhibited activity towards fluorogenic amidomethylcoumarin peptides, but with different substrate preferences. The results show that G. intestinalis contains multiple proteases, many of which are of the cysteine type.
Williams
A G
AG
Department of Zoology, University of Glasgow, UK.
Coombs
G H
GH
D000595:Amino Acid Sequence; D000818:Animals; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D010450:Endopeptidases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D005456:Fluorescent Dyes / Q000737:chemistry; D016829:Giardia lamblia / Q000201:enzymology* / Q000254:growth & development; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009842:Oligopeptides / Q000737:chemistry; D011480:Protease Inhibitors / Q000494:pharmacology; D013379:Substrate Specificity
D005456:Fluorescent Dyes; D009842:Oligopeptides; D011480:Protease Inhibitors; C006164:S 2160; D010450:Endopeptidases; D003546:Cysteine Endopeptidases