Effects of medium viscosity on kinetics of the enzymatic reaction catalyzed by bacterial RNase

Effects of medium viscosity on kinetic parameters of poly(U) hydrolysis cat alyzed by RNase from Bac. intermedius 7P (binase) were studied in solutions of sucrose (4-50 wt. %) and glycerol (35-62 wt. %) in Tris-sodium acetate buffer (pH 7.5) at 25 degrees C. The rate constant of reaction kw, was prac tically unchanged over a wide range of viscosities (1-15 cP for sucrose and 2.5-3 cP for glycerol). In glycerol solutions, k(cat) slightly increased w ith viscosity increase from 4 to 10 cP. Addition of NaCl to the buffer medi um resulted in an inhibitory effect of Na+ on k(cat), prevented by 50% sucr ose or 60% glycerol. It is concluded that binase-catalyzed poly(U) cleavage occurs through a "tense"-substrate mechanism, similarly to reactions catal yzed by alpha-chymotrypsin, trypsin, and laccase.