Ditson S L, Sung S M, Mayer R M
Arch Biochem Biophys. 1986 Aug 15;249(1):53-60. doi: 10.1016/0003-9861(86)90559-x.
Dextransucrase, from Streptococcus sanguis ATCC 10558, was immobilized on hydroxylapatite and was "charged" in short pulses with labeled sucrose, as previously described [V. K. Parnaik, G. A. Luzio, D. A. Grahame, S. L. Ditson, and R. M. Mayer (1983) Carbohydr. Res. 121, 257-268]. The "charged" enzyme has been shown to contain both bound glucose and gluco-oligosaccharides. The reactivity of this form of the enzyme has been studied, and shown to have unexpected behavior. Earlier pulse-chase experiments [J. F. Robyt, B. K. Kimble, and T. F. Walseth (1979) Arch. Biochem. Biophys. 165, 634-640; S. L. Ditson and R. M. Mayer (1984) Carbohydr. Res. 126, 170-175], carried out with high concentrations of unlabeled sucrose in the chase, resulted in a rapid decrease in isotope at the reducing termini of enzyme-bound oligosaccharides. However, in the present work, in which the pulsed enzyme was chased with low concentrations of unlabeled sucrose, we observed an increase in the radioactive reducing termini. The possibility that this was due to the enzymatic hydrolysis of dextran has been ruled out. Data presented demonstrate that the enzyme catalyzes the depolymerization of the bound oligosaccharides. Individual glucosyl residues of the oligosaccharides are transferred to acceptors, such as added maltose to form a trisaccharide, or water to form glucose. Similarly, the glucosyl residues can be transferred to added fructose to form sucrose. The studies also provide evidence that the oligosaccharides are slowly released from the enzyme. The ability of the enzyme to catalyze the reverse of the glucosyl transfer reaction involving acceptors was also examined. It was observed that glucose residues transferred by dextransucrase to an acceptor can also be removed to produce sucrose when fructose is added.
将来自血链球菌ATCC 10558的葡聚糖蔗糖酶固定在羟基磷灰石上，并如先前所述[V. K. 帕尔奈克、G. A. 卢齐奥、D. A. 格雷厄姆、S. L. 迪特森和R. M. 迈耶（1983年）《碳水化合物研究》121卷，257 - 268页]，用标记的蔗糖进行短脉冲“充电”。已证明“充电”后的酶同时含有结合的葡萄糖和葡糖寡糖。对这种形式的酶的反应活性进行了研究，并显示出意想不到的行为。早期的脉冲追踪实验[J. F. 罗比特、B. K. 金布尔和T. F. 瓦尔塞思（1979年）《生物化学与生物物理学报》165卷，634 - 640页；S. L. 迪特森和R. M. 迈耶（1984年）《碳水化合物研究》126卷，170 - 175页]，在追踪过程中使用高浓度的未标记蔗糖，导致酶结合寡糖还原末端的同位素迅速减少。然而，在本研究中，用低浓度的未标记蔗糖对脉冲酶进行追踪时，我们观察到放射性还原末端有所增加。这种情况是由于葡聚糖的酶促水解所致的可能性已被排除。所呈现的数据表明该酶催化结合寡糖的解聚。寡糖的单个葡萄糖基残基被转移到受体上，例如添加的麦芽糖形成三糖，或水形成葡萄糖。同样，葡萄糖基残基也可以转移到添加的果糖上形成蔗糖。这些研究还提供了证据表明寡糖会从酶中缓慢释放。还研究了该酶催化涉及受体的葡糖基转移反应逆反应的能力。观察到葡聚糖蔗糖酶转移到受体上的葡萄糖残基在添加果糖时也可以被去除以产生蔗糖。
