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首页> 外文期刊>Sugar Tech >Production, Partial Purification and Kinetic Characterization of Dextransucrase from Leuconostoc mesenteroides in Relation to Dextran Problem in Sugarcane
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Production, Partial Purification and Kinetic Characterization of Dextransucrase from Leuconostoc mesenteroides in Relation to Dextran Problem in Sugarcane

机译：猪肠球菌葡聚糖葡聚糖酶的生产，部分纯化和动力学特性与甘蔗葡聚糖问题的关系

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Dextransucrase was produced from Leuconostoc mesenteroides strain MTCC 107. Maximum enzyme production was observed after 60 h of incubation, with drop in pH from 7.0 at 0h to 5.5 at 60 h. The crude enzyme was subjected to ammonium sulphate fractionation and DEAE-cellulose anion-exchange chromatography. Three isoforms of dextransucrase viz. DS-Ⅰ, DS-Ⅱ and DS-Ⅲ, each with optimum pH 5.5 and optimum temperature 45 ℃ were identified. Isoforms DS-Ⅰ and DS-Ⅲ were found to be thermostable between 10 and 60 ℃ while isoform DS-Ⅱ was thermostable between 10 and 50 ℃. The K_m values (for sucrose as substrate) of the isoforms DS-Ⅰ, DS-Ⅱ and DS-Ⅲ at their optimum pH and temperature were 206.34, 492.45 and 108.00 mM, respectively. Corresponding V_(max) values were 428.08, 505.58 and 410.85 μmol reducing sugars produced/min/mg protein, respectively. The pK_a values of ionizing groups obtained by using Dixon and Marangoni methods suggested the participation of Asp/Glu residues and His residues in both substrate binding and catalysis. Lineweaver Burk plots obtained by varying concentrations of dextran (as substrate) at fixed sucrose concentration were found to be biphasic for all the three dextransucrase isoforms, suggesting negative cooperativ-ity. Both the transferase and sucrase activities of the dextransucrase isoforms were markedly inhibited by sodium lauryl sulphate, barium chloride, EDTA, potassium chloride, manganese chloride and sodium metasilicate. These metal salts/chemicals when exogenously added to sugarcane juice effectively reduced the dextran formation during storage.
机译：葡聚糖转化酶是由肠膜明串珠菌MTCC 107菌株产生的。孵育60小时后观察到最大的酶产生，pH从0小时的7.0降至60小时的5.5。将粗酶进行硫酸铵分级分离和DEAE-纤维素阴离子交换色谱。葡糖转移酶的三种同工型。分别确定了最适pH 5.5和最适温度45℃的DS-Ⅰ，DS-Ⅱ和DS-Ⅲ。发现亚型DS-Ⅰ和DS-Ⅲ在10至60℃之间是热稳定的，而亚型DS-Ⅱ在10至50℃之间是热稳定的。亚型DS-Ⅰ，DS-Ⅱ和DS-Ⅲ在其最佳pH和温度下的K_m值（以蔗糖为底物）分别为206.34、492.45和108.00 mM。相应的V_（max）值分别为428.08、505.58和410.85μmol产生的还原糖/ min / mg蛋白。通过Dixon和Marangoni方法获得的电离基团的pK_a值表明，Asp / Glu残基和His残基参与底物结合和催化。发现在固定的蔗糖浓度下通过改变右旋糖酐（作为底物）的浓度获得的Lineweaver Burk图对所有三种右旋糖酐酶同工型都是两相的，表明其负合作性。月桂基硫酸钠，氯化钡，EDTA，氯化钾，氯化锰和偏硅酸钠显着抑制了葡聚糖转氨酶同工型的转移酶和蔗糖酶活性。这些金属盐/化学品在外源添加到甘蔗汁中时，可以有效地减少储存过程中葡聚糖的形成。

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