Wang Fei, Zhang Yu-Qing
Silk Biotechnology Laboratory, School of Biological and Basic Medical Sciences, Soochow University, Suzhou, PR China.
Silk Biotechnology Laboratory, School of Biological and Basic Medical Sciences, Soochow University, Suzhou, PR China.
Adv Protein Chem Struct Biol. 2015;98:263-91. doi: 10.1016/bs.apcsb.2014.11.005. Epub 2015 Mar 3.
Bombyx mori silk fibroin is a type of protein-based polymer with unique characteristics that is widely used in the research and development of medical biomaterials. The degummed filament of silk fibroin can be dissolved in a highly concentrated salt solution. After desalination, the regenerated liquid silk fibroin (LSF) solution could be made into various forms of silk biomaterials, such as powder, fiber, film, porous matrix, 3D scaffold, and hydrogel, depending on its application. In this study, we mixed the liquid silk solution with enzymes, including oxidase and hydrolase, and rapidly injected the mixture into an excess of acetone. The enzyme retained most of its enzymatic activity and was also captured in silk fibroin nanoparticles (SFNs), which instantly formed via a configuration transition of the regenerated silk protein from a random coil and α-helix to a β-sheet. The resulting enzyme-captured SFNs displayed a fine crystal structure with a high activity recovery and good thermal stability. Moreover, the affinities of these modified enzymes to their substrate did not evidently suffer from the capture. When only the liquid silk solution was rapidly injected into acetone, the resulting globular SFNs with the same crystallinity were also a good carrier that was covalently conjugated to enzymes and insulin. Thus, silk protein nanoparticles are of potential value as an enzyme or peptide delivery system for the research and development of medical biomaterials. In this report, the bioconjugation of SFNs with glucose oxidase, superoxidase, β-glucosidase, L-asparaginase, neutral protease, and insulin and their characterization are described in detail.
家蚕丝素蛋白是一种具有独特特性的基于蛋白质的聚合物，广泛应用于医用生物材料的研发。脱胶的丝素蛋白长丝可溶解于高浓度盐溶液中。脱盐后，再生液体丝素蛋白（LSF）溶液可根据其应用制成各种形式的丝生物材料，如粉末、纤维、薄膜、多孔基质、3D支架和水凝胶。在本研究中，我们将液体丝溶液与包括氧化酶和水解酶在内的酶混合，并将混合物迅速注入过量的丙酮中。酶保留了大部分酶活性，并且还被捕获在丝素蛋白纳米颗粒（SFN）中，这些纳米颗粒通过再生丝蛋白从无规卷曲和α-螺旋到β-折叠的构象转变立即形成。所得的酶捕获的SFN显示出具有高活性回收率和良好热稳定性的精细晶体结构。此外，这些修饰酶与其底物的亲和力在捕获后并未明显受损。当仅将液体丝溶液迅速注入丙酮中时，所得具有相同结晶度的球形SFN也是一种与酶和胰岛素共价偶联的良好载体。因此，丝蛋白纳米颗粒作为医用生物材料研发的酶或肽递送系统具有潜在价值。在本报告中，详细描述了SFN与葡萄糖氧化酶、超氧化物酶、β-葡萄糖苷酶、L-天冬酰胺酶、中性蛋白酶和胰岛素的生物偶联及其表征。
