Király Márton, Kiss Borbála Dalmadi, Horváth Péter, Drahos László, Mirzahosseini Arash, Pálfy Gyula, Antal István, Ludányi Krisztina
Department of Pharmaceutics, Faculty of Pharmacy, Semmelweis University, Hőgyes Endre u. 7., 1092, Budapest, Hungary.
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Semmelweis University, Hőgyes Endre u. 7., 1092, Budapest, Hungary.
Biotechnol Rep (Amst). 2021 May 26;30:e00637. doi: 10.1016/j.btre.2021.e00637. eCollection 2021 Jun.
Thermal stability of lactase (β-galactosidase) enzyme has been studied by a variety of physico-chemical methods. β-galactosidase is the main active ingredient of medications for lactose intolerance. It is typically produced industrially by the Aspergillus oryzae filamentous fungus. Lactase was used as a model to help understand thermal stability of enzyme-type biopharmaceuticals. Enzyme activity (hydrolyzation of lactose) of β-galactosidase was determined after storing the solid enzyme substance at various temperatures. For a better understanding of the relationship between structure and activity changes we determined the mass and size of the molecules with gel electrophoresis and dynamic light scattering and detected aggregation processes. A bottom-up proteomic procedure was used to determine the primary amino acid sequence and to investigate changes in the N-glycosylation pattern of the protein. NMR and CD spectroscopic methods were used to observe changes in higher order structures and to reveal relationships between structural and functional changes.
已通过多种物理化学方法研究了乳糖酶（β-半乳糖苷酶）的热稳定性。β-半乳糖苷酶是治疗乳糖不耐受药物的主要活性成分。它通常由米曲霉丝状真菌在工业上生产。乳糖酶被用作模型来帮助理解酶型生物药物的热稳定性。在将固体酶物质储存在不同温度下后，测定了β-半乳糖苷酶的酶活性（乳糖水解）。为了更好地理解结构与活性变化之间的关系，我们用凝胶电泳和动态光散射测定了分子的质量和大小，并检测了聚集过程。采用自下而上的蛋白质组学方法来确定一级氨基酸序列，并研究蛋白质N-糖基化模式的变化。使用核磁共振和圆二色光谱方法来观察高级结构的变化，并揭示结构与功能变化之间的关系。
