百合鳞茎蔗糖合成酶活性检测体系的建立

Establishment of Detection System for Sucrose Synthase Activity in Lily Bulb

为了建立富含多糖的百合鳞茎蔗糖合成酶(sucrose synthase,EC2.4.1.13,SuSy)活性检测体系,深入研究其蔗糖代谢机制,以兰州百合(Lilium davidii var.unicolor)鳞茎外层鳞片为试材,分别研究了提取缓冲液种类及pH值、反应温度、底物浓度以及缓冲液pH值对SuSy合成和分解方向活性的影响。结果表明:SuSy合成方向活性检测的最适提取缓冲液是pH值为7.8的Tris-HCl,最适反应温度为50℃,底物果糖最适浓度为50 mmol.L-1,UDPG最适浓度为5 mmol.L-1,反应缓冲液Tris-HCl最适pH值为7.5;SuSy分解方向活性检测的最适提取缓冲液为pH值7.8的Hepes-NaOH,最适反应温度为40℃,底物蔗糖最适浓度为10mmol.L-1,UDP最适浓度为7 mmol.L-1,反应缓冲液Mes-NaOH最适pH值为4.5。

This investigation was designed to establish the detection system for sucrose synthase （EC 2.4.1.13, SuSy） activities inlily bulb enriched with polysaccharides, which provided a detection method for the further research on the mechanism of sucrose metabolism. The effects of extracting buffer types, pH, reaction temperature, substrate concentrations, pH of the reaction buffer on the SuSy activities in both synthesis and decomposition direction were respectively studied by using the exterior scales of Lilium davidii var. unicolor at planting stage as materials. And the results showed that the optimum extraction buffer was Tris-HC1 of pH7.8, the appropriate temperature was 50 ℃, the preferential substrate concentration of fructose was 50 mmol ·L^-1, the preferential substrate concentration of UDPG was 5 mmol·L^-1, and the suitable pH for reaction buffer Tris-HC1 was 7.5 in the detection of SuSy synthesis activities. In the detection of SuSy decomposition activities, the optimum extraction buffer was HepesNaOH of pH7.8, the appropriate temperature was 40℃, the adequate substrate concentration of sucrose was 10 mmol·L^-1, the adequate substrate concentration of UDP was 7 mmol·L^-1, and the suitable pH for reaction buffer Mes-NaOH was 4.5.