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首页> 外文期刊>Journal of Polymers and the Environment >A Novel Affinity Chromatographic Material for the Purification of Extracellular Polyhydroxybutyrate Depolymerases
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A Novel Affinity Chromatographic Material for the Purification of Extracellular Polyhydroxybutyrate Depolymerases

机译：一种新型亲和层析材料，用于纯化细胞外多羟基丁酸酯解聚酶

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摘要

A novel affinity chromatographic material, which is composed of silica matrix, coated with poly hydroxybutyrate (PHB) powder, suitable for the purifica tion of PHB depolymerases, was developed. The surface morphology of the PHB-silica coated particles (silica-PHB composite particles) was examined by scanning electron microscopy and revealed a successful uniform coating of silica particles with PHB. Moreover, the complex of these materials retained its homogeneity even after incubation at 80 °C for 6 h, whereas the strong binding of PHB on silica surface was further verified by thermal gravimetric analysis and by PHB extraction- from silica surface- experiments. This novel material was demonstrated to be suitable for both, the one-step on-batch and on-column purification of Thermus thermophilus extracellular PHB depolymerase. The enzyme exhibited higher affinity against the composite of silica-PHB particles than PHB powder, since the one step purification-fold and the overall recovery of the enzyme were 2.8 and 4 times higher respectively, in the first case. Reusability of the silica-PHB composites parti cles was examined by determining the recoveries of PHB depolymerase. The enzyme recoveries were ranged from 30 to 35% for the first five uses, whereas for further uses recoveries gradually dropped to 15-18% indicating that the particles could be used repeatedly for five times. This material could be also a suitable support for lipases or other proteins that exhibit strong affinity to hydrophobic materials.
机译：开发了一种新型的亲和色谱材料，该材料由二氧化硅基质组成，并涂有聚羟基丁酸酯（PHB）粉末，适用于纯化PHB解聚酶。通过扫描电子显微镜检查了PHB-二氧化硅涂覆的颗粒（二氧化硅-PHB复合颗粒）的表面形态，并显示了用PHB成功地均匀涂覆了二氧化硅颗粒。此外，即使在80°C下孵育6小时后，这些材料的复合物仍保持其均一性，而通过热重分析和PHB提取（从二氧化硅表面实验）进一步证实了PHB在二氧化硅表面上的牢固结合。事实证明，这种新型材料适用于嗜热栖热菌细胞外PHB解聚酶的单步批量纯化和柱上纯化。该酶对二氧化硅-PHB颗粒的复合物表现出比PHB粉末更高的亲和力，因为在第一种情况下，一步纯化倍数和酶的总回收率分别高2.8和4倍。通过确定PHB解聚酶的回收率来检查二氧化硅-PHB复合材料颗粒的可重用性。前五次使用的酶回收率范围为30％至35％，而其他用途的回收率逐渐下降至15-18％，表明该颗粒可重复使用五次。该材料也可以是脂肪酶或其他对疏水性材料具有强亲和力的蛋白质的合适支持物。

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