体外酶法合成的糖原状聚合物在生物、医疗、化工等领域具有广泛的应用,其较小的分子尺寸(106 Da及之下)使其具有良好的应用效果。该研究通过蔗糖磷酸化酶-α葡聚糖磷酸化酶-分支酶-4-α糖基转移酶的四酶协同法合成出分子质量为106 Da的糖原状聚合物,通过控制酶活比例制备了一系列具有不同分子结构的糖原状聚合物,并分别通过高效凝胶尺寸排阻色谱,高效阴离子交换色谱和核磁共振氢谱等手段对糖原状聚合物的分子尺寸和精细结构进行表征。结果表明,糖原状聚合物的分子质量成功从2.20×107 Da降至4.94×106 Da并最终稳定在106 Da;平均链长在8.11~9.99,分支度保持在7.4%~10.7%,保留了高度分支化的结构,粒径Rz在19.3~28.0 nm,抗酶解率在39.78%~43.19%;为糖原状聚合物的体外酶法合成提供了新思路,也增加了在生物医疗等领域的应用潜力。
Glycogen-like- polymer is synthesized by biological enzyme method in vitro and is widely used in biology, medical treatment, chemical industry and other fields.Its small molecular size (the molecular weight is 106 Da and below) makes it have good application effect.In this study, glycogen-like- polymer with molecular weight of 106 Da was synthesized by the synergistic method of sucrose phosphorylase-α-glucan phosphorylase-branching enzyme-4-α-Glycosyltransferase.A series of glycogen-like- polymer samples with different molecular sizes and fine structures were synthesized by controlling the ratio of enzyme activities of reaction elements.The high-performance gel size exclusion chromatography, high-performance anion chromatography and nuclear magnetic resonance hydrogen spectrum were used respectively to characterize the molecular size and fine structure.The results showed that the molecular weight of Glycogen-like- polymer reduce from 2.20×107 Da to 4.94×106 Da successfully and finally stabilize at 106 Da.The average chain length is between 8.11 and 9.99.The branching degree remained at 7.4%-10.7% showed it retain the highly branched structure.The particle size Rz was between 19.3 nm and 28.0 nm and the anti-enzymatic hydrolysis rate was between 39.78% and 43.19%.This provides a new idea for the in vitro enzymatic synthesis of glycogen-like-polymer, and also increases the application potential in biomedical fields.
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