Isolation and Characterization of Proteinases from the Sarcocarp of Snake-Gourd Fruit

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Abstract
Seven proteinases were isolated from the fruit of snake-gourd, Trichosanthes cucumeroides Maxim . Their isozymes are all serine proteinases, and homologous in their respective molecular weights, amino acid compositions, and enzymatic properties. Their molecular weight was estimated to be about 50,000. Using casein as a substrate, the maximum activity was found in the alkaline pH region. The optimum temperature using casein was 70°C at pH 7.3. The enzymes were strongly inhibited by diisopropyl fluorophosphate and not inhibited by inhibitors of sulfhydryl or metalloproteases. The reduced and S -carboxyniethylated insulin B-chain was used as a substrate in an investigation of the specificity. The enzyme was found to have a wide spacificity for this substrate but preferentially hydrolyzed the peptide bonds involving the carboxyl groups of charged amino acid such as S -cm-cysteine, glutamic acid, histidine, arginine, and lysine. Experimental evidence indicated that the snake-gourd proteinases are similar in their properties to cucumisin, which is isolated from the sarcocarp of melon fruit.