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Báo cáo khoa học: A kinetic study of sugarcane sucrose synthase

The kinetic data on sugarcane (Saccharumspp. hybrids) sucrose synthase (SuSy, UDP-glucose: D-fructose 2-a-D-glucosyltransferase, EC 2.4.1.13) are limited. We character-ized kinetically a SuSy activity partially purified from sugarcane variety N19 leaf roll tissue. Primary plot analysis and product inhibition studies showed that a compulsory order ternary complex mechanism is followed, with UDP binding first and UDP-glucose dissociating last from the enzyme. | Eur. J. Biochem. 271 3971-3977 2004 FEBS 2004 doi 10.1111 j.1432-1033.2004.04288.x A kinetic study of sugarcane sucrose synthase Wolfgang E. Schafer1 Johann M. Rohwer2 and Frederik C. Botha3 institute for Plant Biotechnology and department of Biochemistry University of Stellenbosch South Africa 3South African Sugarcane Research Institute Mount Edgecombe South Africa The kinetic data on sugarcane Saccharum spp. hybrids sucrose synthase SuSy UDP-glucose D-fructose 2-a-D-glucosyltransferase EC 2.4.1.13 are limited. We characterized kinetically a SuSy activity partially purified from sugarcane variety N19 leaf roll tissue. Primary plot analysis and product inhibition studies showed that a compulsory order ternary complex mechanism is followed with UDP binding first and UDP-glucose dissociating last from the enzyme. Product inhibition studies showed that UDP-glu-cose is a competitive inhibitor with respect to UDP and a mixed inhibitor with respect to sucrose. Fructose is a mixed inhibitor with regard to both sucrose and UDP. Kinetic constants are as follows Km values mM SE were for sucrose 35.9 2.3 for UdP 0.00191 0.00019 for UDP-glucose 0.234 0.025 and for fructose 6.49 0.61. KS values were for sucrose 227 mM for UDP 0.086 mM for UDP-glucose 0.104 and for fructose 2.23 mM. Replacing estimated kinetic parameters of SuSy in a kinetic model of sucrose accumulation with experimentally determined parameters of the partially purified isoform had significant effects on model outputs with a 41 increase in sucrose concentration and 7.5-fold reduction in fructose the most notable. Of the metabolites included in the model fructose concentration was most affected by changes in SuSy activity doubling and halving of SuSy activity reduced and increased the steady-state fructose concentration by about 42 and 140 respectively. It is concluded that different isoforms of SuSy could have significant differential effects on metabolite concentrations in vivo therefore impacting on metabolic