Study on a proteolytic enzyme from Trypanosoma congolense. Purification and some biochemical properties.
A protease has been purified from Trypanosoma congolense bloodstream forms by osmotic disruption, freeze-thawing of the cells, followed by chromatography using Thiopropyl-Sepharose and gel filtration. The enzyme is a thiolprotease. A combination of SDS-polyacrylamide gel electrophoresis and contact print zymograms using casein as substrate showed a single proteolytic band with a molecular weight of 31 000. The isoelectric point of the enzyme as ascertained by isoelectric focusing extended from pH 4.4 to 5.5 with a maximum at pH 5.0. The protease cleaved various heat denatured substrates such as casein, hemoglobin, albumin and ovalbumin. The highest enzyme activity was observed at pH 5.5 and pH 6.0 using casein and hemoglobin as substrates respectively. The max. temperature was found to be 50 degrees C. The enzyme is inactivated by mercurial compounds, iodoacetamide, iodoactate, chloromethylketones and leupeptin and is activated by dithioerythritol.
D000818:Animals; D002364:Caseins / Q000378:metabolism; D002847:Chromatography, Agarose; D003546:Cysteine Endopeptidases; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000302:isolation & purification* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D011480:Protease Inhibitors; D013379:Substrate Specificity; D014345:Trypanosoma / Q000201:enzymology*
D002364:Caseins; D011480:Protease Inhibitors; D010450:Endopeptidases; D003546:Cysteine Endopeptidases