Cysteine protease from the malaria parasite,Plasmodium berghei-purification and biochemical characterization [Malarya paraziti Plasmodium berghei'den sistein proteaz- saflaşti{dotless}ri{dotless}lmas{dotless} ve karakterizasyonu]

Plasmodium berghei was isolated from mice red blood cells and phase-separated by triton x-100 temperature-induced phase separation procedures. the enzyme cysteine protease was purified 5.33 fold with a recovery of 58%. sds-page analysis of the enzyme revealed two protein bands with molecular weights corresponding to 18 and 40 kda,respectively. the enzyme was optimally active at temperature of 40°c and at a ph of 5.0 (50 mm acetate buffer). activation energy (6.27 kj/mole) of the enzyme was determined from arrhenius plots and initial velocity studies revealed km and vmax values of 2.5 mg/ml and 0.2 μmol/min,respectively. the enzyme was inactive on the substrates,albumin and myoglobin. the enzyme was exclusively sensitive to the cysteine protease specific inhibitor iodoacetate (iaa),but was insensitive to phenylmethylsulphonyl chloride (pmsf),1,10 phenanthroline,soybean trypsin inhibitor (sbti),pepstatin a and edta. the synthetic compounds pp-54 and pp-56,currently being evaluated for their anti-malaria potential,competitively inhibited the enzyme activity with corresponding ki values of 48.88 μg/ml and 0.14 μg/ml,respectively.