School of Chemical Engineering, Hebei University of Technology, Tianjin 300130, China.
Enzyme Microb Technol. 2011 Jun 10;49(1):11-6. doi: 10.1016/j.enzmictec.2011.04.007. Epub 2011 Apr 15.
Nano-materials have been applied in many fields due to their excellent characteristics, such as the high surface area-to-volume ratio, excellent physicochemical properties and biological compatibility. In this study, multi-walled carbon nanotubes (MWCNTs) were utilized to prepare MWCNTs-papain bioconjugates and then realized the immobilization of papain. MWCNTs functionalized with carboxyl- and amine- groups on their surface were used as immobilization carriers. The immobilization of papain on the functionalized MWCNTs through physical absorption was examined. The conjugates were denoted as MWCNTs-papain bioconjugates. To improve the stability, the bioconjugates were further coated by silica through the biomimetic silicification process that induced by papain (denoted as silica-coated bioconjugates). The as-prepared MWCNTs-papain bioconjugates and the silica-coated bioconjugates were characterized by scanning electron microscopy (SEM) and Fourier transform infrared (FTIR) spectroscopy. The preliminary results showed that the bioconjugates could retain most of the initial activity of papain. Compared to free papain and MWCNTs-papain bioconjugates, the silica-coated bioconjugates exhibited significantly improved thermal, pH and recycling stability. Comparisons of the kinetic parameters between MWCNTs-papain bioconjugates and the silica-coated bioconjugates revealed that the K(m) value of the immobilized papain experienced a slight increase after silica coating, which suggested that the silica coating did not significantly hinder papain's access to substrate or release of product.
纳米材料由于其优异的特性，如高的表面积与体积比、优异的物理化学性质和生物相容性，已被应用于许多领域。在本研究中，多壁碳纳米管（MWCNTs）被用于制备 MWCNTs-木瓜蛋白酶生物缀合物，然后实现了木瓜蛋白酶的固定化。MWCNTs 表面功能化的羧基和氨基被用作固定化载体。通过物理吸附考察了木瓜蛋白酶在功能化 MWCNTs 上的固定化。将固定化的木瓜蛋白酶称为 MWCNTs-木瓜蛋白酶生物缀合物。为了提高稳定性，通过木瓜蛋白酶诱导的仿生硅化过程进一步用硅酸钠对生物缀合物进行涂层（表示为硅涂层生物缀合物）。采用扫描电子显微镜（SEM）和傅里叶变换红外光谱（FTIR）对所制备的 MWCNTs-木瓜蛋白酶生物缀合物和硅涂层生物缀合物进行了表征。初步结果表明，生物缀合物可以保留木瓜蛋白酶的大部分初始活性。与游离木瓜蛋白酶和 MWCNTs-木瓜蛋白酶生物缀合物相比，硅涂层生物缀合物表现出显著提高的热稳定性、pH 值稳定性和循环稳定性。MWCNTs-木瓜蛋白酶生物缀合物和硅涂层生物缀合物的动力学参数比较表明，硅涂层后固定化木瓜蛋白酶的 K(m)值略有增加，这表明硅涂层并没有显著阻碍木瓜蛋白酶与底物的接触或产物的释放。
