ACTIVITY OF PROTEASE EXTRACTED FROM RICE-RHIZOSPHERE SOILS UNDER DOUBLE CROPPING OF RICE AND WHEAT

Protease-active extracts were prepared from 2 rice-rhizosphere and non -rhizosphere soil samples 3 proteolytic isolates from the soil samples which were taken from water logged paddy fields (Gray Lowland Soils) under double cropping of rice and wheat that had been treated with che mical fertilizer (CF) or organic manure (OM). The activities were assa yed and characterized using an artificial substrate, nzyloxycarbonyl-L -phenylalanyl-L-tyrosyl-L-leucine (ZFTL). The protease-active extracts from the rhizosphere and non-rhizosphere of the CF and OM soils (abbr eviated as CFR, CFN, OMR, and OMN) showed an optimum pH between 7 and 8. The extracts hydrolyzed ZFTL to Tyr-Leu and Leu; generally, Tyr-Leu was the main reaction product. Activities of all the samples were inh ibited by EDTA and p-chloromercuribenzoic acid (PCMB) but not by pepst atin and phenylmethanesulfonyl fluoride (PMSF). These results indicate that the main components of the ZFTL-hydrolases in CFR, CFN, OMR, and OMN were essentially similar. Optimum pH for the activity of the prot ease extracted from a Gram negative rod (MNB003) and two actinomycetou s isolates (MNA014 and MRA017) was about 8, 8, and 8.5, respectively. Extract from the culture of MNB003 hydrolyzed ZFTL to Tyr-Leu and Leu, and the protease activity was inhibited by EDTA and PCMB. Protease ac tivities of the extracts from cultures of the two actinomycetes were i nhibited by EDTA but not by PCMB. The extract from culture of MNA014 p roduced mainly Tyr-Leu from ZFTL while MRA017 produced Leu. Characteri stics of the protease activity from cultures of MNB003 and MNA014 were similar to those of the enzyme extracted from a paddy field soil unde r single crapping (Takeuchi and Hayano 1994: Soil Sci. Plant Nutr., 40 , 691-695) and from an Andosol under tomato cultivation (Hayano et al. 1987: Biol. Fertil. Soils, 4, 179-183), respectively.