serine protease from germinated wheat seeds

ABSTRACT

A simple method for the purification of a serine endoprotease from wheat Triticum aestivum [cv. Giza 164] has been developed. It consists of ion-exchange and gel filtration chromatography. The molecular mass of the enzyme was 58 kDa by SDS/PAGE under reducing conditions and 57 kDa by gel filtration on a Sepharose 6B column. The enzyme had isoelectric point and pH optimum at 4.2 and 4.5, respectively. The substrate specificity of the enzyme was studied by the use of synthesized and natural substrates, azocasein, azoalbumin, hemoglobin, casein, gelatin and egg albumin. The enzyme appears to prefer azocasein with a Km of 2 mg azocasein/ml. The enzyme had a temperature optimum at 50°C with heat stability up to 40°C. While Co[2+] and Mg[2+] accelerated the enzyme activity by 54% and 56%, respectively, Ca[2+] and Ni[2+] had very little effect. The enzyme was strongly inhibited by PMSF [phenylmethylsulphonyl fluoride], but not by the other protease inhibitors, suggesting that the enzyme is a serine protease. From these results it can be concluded that the T. aestivum serine protease may be suitable for food processing