Ilesanmi Olutosin Samuel, Adewale Isaac Olusanjo
Department of Chemical Sciences, Achievers University, Owo, Ondo State, Nigeria.
Department of Biochemistry and Molecular Biology, Obafemi Awolowo University, Ile-Ife, Nigeria.
Biotechnol Rep (Amst). 2020 Jul 1;27:e00499. doi: 10.1016/j.btre.2020.e00499. eCollection 2020 Sep.
A shortened method of purification and immobilization of tyrosinase from different species of yam () on insoluble supports is described. The enzyme was purified by aqueous two-phase partitioning (ATPS) followed by gel filtration chromatography. The purified enzyme was immobilized on Ca-alginate, polyacrylamide gel or as cross-linked enzyme aggregate (CLEA) to obtain a yield of between 51-64%, 33-46% and 52-65% respectively for all the yam species. The optimum pH obtained for tyrosinase immobilized on polyacrylamide gel and CLEA was equivalent to that of free enzyme (pH 6.5). In contrast, Ca-alginate entrapped tyrosinase exhibited a shift of optimum pH to 7.0. Entrapped Tyrosinase in polyacrylamide gel and Ca-alginate also retained the same optimum temperature as the free enzyme (50 °C). While the optimum temperature of CLEA shifted to 60 °C. When subjected to four repeated use cycles, tyrosinase entrapped in polyacrylamide gel, Ca-alginate and CLEA still retained close to 40, 35 and 45 % of their initial activities respectively after the fourth cycle. The overall result further suggests yam tyrosinase as a promising enzyme for biocatalysis and biotechnological applications.
本文描述了一种从不同种类山药中纯化和固定化酪氨酸酶到不溶性载体上的简化方法。该酶先通过双水相分配（ATPS）进行纯化，然后进行凝胶过滤色谱。将纯化后的酶固定在海藻酸钙、聚丙烯酰胺凝胶上或制成交联酶聚集体（CLEA），所有山药种类的固定化产率分别在51 - 64%、33 - 46%和52 - 65%之间。固定在聚丙烯酰胺凝胶和CLEA上的酪氨酸酶的最适pH与游离酶相同（pH 6.5）。相比之下，海藻酸钙包埋的酪氨酸酶的最适pH向7.0偏移。聚丙烯酰胺凝胶和海藻酸钙中包埋的酪氨酸酶也保留了与游离酶相同的最适温度（50°C）。而CLEA的最适温度则偏移到了60°C。当进行四个重复使用周期时，聚丙烯酰胺凝胶、海藻酸钙和CLEA中包埋的酪氨酸酶在第四个周期后仍分别保留其初始活性的近40%、35%和45%。总体结果进一步表明，山药酪氨酸酶是一种有前途的用于生物催化和生物技术应用的酶。
