Kinetic properties of raffinose synthase from rice (Oryza sativa L.)

The kinetic properties of purified recombinant rice raffinose synthase (RS) suggested the catalytic mechanisms for RS for both hydrolysis and synthesis reactions. The hydrolysis Km values of rice RS were found to be 2.15, 10.9 and 12.0 mM for galactinol, raffinose, and p-nitrophenyl-α-d-galactopyranoside, respectively. In synthesis reactions, the enzyme transferred galactose from galactinol to sucrose to yield raffinose and myo-inostiol. The raffinose synthesis was best fitted to a Ping-Pong Bi-Bi mechanism, with Km values of 6.7 and 18.5 mM estimated at non-saturated substrate levels for galactinol and sucrose. The synthesis, catalyzed by rice RS was reversible, the forward reactions approached equilibrium after 4 h, and the Keq was calculated to be 4.46. The specific activity 25.6 nkat/mg for raffinose formation was more efficient than that of other RS.