a Department of Chemistry, Faculty of Science & Arts , Inonu University , Malatya , Turkey.
Artif Cells Nanomed Biotechnol. 2018;46(sup2):1035-1045. doi: 10.1080/21691401.2018.1478422. Epub 2018 Jun 6.
l-Asparaginase (l-ASNase) is a vital enzyme for medical treatment and food industry. Here, we assessed the use of FeO@Mobil Composition of Matter No. 41 (MCM-41) magnetic nanoparticles as carrier matrix for l-ASNase immobilization. In addition, surface of FeO@MCM-41 magnetic nanoparticles was functionalized with 3-mercaptopropyltrimethoxysilane (MPTMS) to enhance stability of l-ASNase. The chemical structure, thermal properties, magnetic profile and morphology of the thiol-functionalized FeO@MCM-41 magnetic nanoparticles were characterized with Fourier transform infrared spectroscopy (FTIR), thermogravimetric analysis (TGA), differential thermal analysis (DTA), differential scanning calorimetry (DSC), vibrating sample magnetometer (VSM), scanning electron microscope (SEM), energy dispersive X-ray (EDX) spectroscopy and zeta-potential measurement. l-ASNase was covalently immobilized onto the thiol-functionalized FeO@MCM-41 magnetic nanoparticles. The properties of the immobilized enzyme, including optimum pH, temperature, kinetic parameters, thermal stability, reusability and storage stability were investigated and compared to free one. Immobilized enzyme was found to be stable over a wide range of pH and temperature range than free enzyme. The immobilized l-ASNase also showed higher thermal stability after 180 min incubation at 50 °C. The immobilized enzyme still retained 63% of its original activity after 16 times of reuse. The Km value for the immobilized enzyme was 1.15-fold lower than the free enzyme, which indicates increased affinity for the substrate. Additionally, the immobilized enzyme was active over 65% and 53% after 30 days of storage at 4 °C and room temperature (∼25 °C), respectively. Thereby, the results confirmed that thiol-functionalized FeO@MCM-41 magnetic nanoparticles had high efficiency for l-ASNase immobilization and improved stability of L-ASNase.
L-天冬酰胺酶（l-ASNase）是医学治疗和食品工业的重要酶。在这里，我们评估了使用 FeO@ Mobil 组成物质号 41（MCM-41）磁性纳米粒子作为 l-ASNase 固定化的载体基质。此外，FeO@MCM-41 磁性纳米粒子的表面用 3-巯基丙基三甲氧基硅烷（MPTMS）进行功能化，以提高 l-ASNase 的稳定性。巯基功能化的 FeO@MCM-41 磁性纳米粒子的化学结构、热性能、磁性轮廓和形态用傅里叶变换红外光谱（FTIR）、热重分析（TGA）、差示热分析（DTA）、差示扫描量热法（DSC）、振动样品磁强计（VSM）、扫描电子显微镜（SEM）、能量色散 X 射线（EDX）光谱和 ζ-电位测量来表征。l-ASNase 被共价固定在巯基功能化的 FeO@MCM-41 磁性纳米粒子上。研究了固定化酶的性质，包括最适 pH、温度、动力学参数、热稳定性、可重复使用性和储存稳定性，并与游离酶进行了比较。与游离酶相比，固定化酶在较宽的 pH 和温度范围内更稳定。固定化 l-ASNase 在 50°C 孵育 180 分钟后也表现出更高的热稳定性。固定化酶在重复使用 16 次后仍保留其原始活性的 63%。固定化酶的 Km 值比游离酶低 1.15 倍，这表明其对底物的亲和力增加。此外，固定化酶在 4°C 和室温（约 25°C）下储存 30 天后的活性分别为 65%和 53%。因此，结果证实巯基功能化的 FeO@MCM-41 磁性纳米粒子对 l-ASNase 固定化具有高效率，并提高了 L-ASNase 的稳定性。
