Extracellular ribonuclease from Rhizopus stolonifer: characteristics of an atypical--guanylic acid preferential--enzyme from ribonuclease T2 family.

An extracellular ribonuclease from Rhizopus stolonifer (designated as RNase Rs) was purified to homogeneity by chromatography on DEAE-cellulose followed by CM-cellulose. The Mr of the purified enzyme determined by gel filtration and SDS-PAGE is 25,000 and 28,200, respectively. RNase Rs is a glycoprotein and contains 10.5% neutral sugar. It is an acidic protein with a pI of 5.0 and has a blocked N-terminus. The optimum pH and temperature are 5.5 and 45 degrees C, respectively. RNase Rs shows high stability between pH 6.0-10.0. Divalent cations like Zn2+, Hg2+ and Cu2+ inhibit the enzyme activity whereas, mononucleotides does not have any significant effect. The enzyme cleaves RNA to 3'-mononucleotides via 2',3'-cyclic nucleotides, with preferential liberation of 2',3'-cyclic GMP, suggesting that RNase Rs is a guanylic acid preferential cyclizing RNase. Moreover, cyclic nucleotides generated are highly resistant to further hydrolysis.