Purification and characterization of a highly selective sucrose isomerase from Erwinia rhapontici NX-5

摘要

A highly selective sucrose isomerase (Slase) was purified to homogeneity from the cell-free extract of Erwinia rhapontici NX-5 with a recovery of 27.7% and a fold purification of 213.6. The purified Slase showed a high specific activity of 427.1 U mg~(-1) with molecular weight of 65.6 kDa. The K_m for sucrose was 222 mM while V_(max) was 546 U mg~(-1). The optimum pH and temperature for Slase activity were 6.0 and 30 ℃, respectively. The purified Slase was stable in the temperature range of 10-40 °C and retained 65% of the enzyme activity after 2 weeks' storage at 30 ℃. The Slase activity was enhanced by Mg~(2+) and Mn~(2+), inhibited by Ca~(2+), Cu~(2+), Zn~(2+), and Co~(2+), completely inhibited by Hg~(2+) and Ag~(2+). The purified Slase was strongly inhibited by SDS, while partially inhibited by dimethylformamide, tetrahydrofuran, and PMSF. Additionally, glucose and fructose acted as competitive inhibitors for purified Slase.