Functionalized Graphene Oxide with Chitosan for Protein Nanocarriers to Protect against Enzymatic Cleavage and Retain Collagenase Activity.
Functionalized Graphene Oxide with Chitosan for Protein Nanocarriers to Protect against Enzymatic Cleavage and Retain Collagenase Activity.
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DOI：
10.1038/srep42258
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发表时间：
2017-02-10
影响因子：
4.6
通讯作者：
Ghasemi Y
中科院分区：
文献类型：
--
作者：
Emadi F;Amini A;Gholami A;Ghasemi Y
由于酶的切割，蛋白质的半衰期很短。本文提出了一种由氧化石墨烯(GO) + 壳聚糖(CS)制成的新型蛋白质纳米载体，在成功阻止蛋白质降解的同时保持了蛋白质的活性。将牛血清白蛋白(BSA)和胶原酶负载于GO和GO-CS上，考察蛋白质的稳定性和活性。采用扫描电子显微镜、原子力显微镜、透射电子显微镜、差示扫描量热仪、紫外-可见光谱、傅立叶变换红外光谱、背散射光谱、拉曼光谱、十二烷基硫酸钠-聚丙烯酰胺凝胶和酶谱等技术对其进行了表征。以GO-BSA和GO-CS-BSA为原料，在37 °C条件下，分别加入GO-CS-BSA和GO-CS-BSA，在37 ℃下消化0.5h-1-3-6h，通过对牛血清白蛋白的酶切分析，探讨GO和GO-CS-BSA的保护作用。表征表明，成功地合成了几层GO，并被CS覆盖。根据明胶酶谱分析，GO和GO-CS上负载的胶原酶裂解明胶并产生无染色的条带，证实了负载胶原酶的活性。SDS-PAGE分析显示，GO-BSA和GO-CS-BSA处理30min和1h后，GO-BSA和GO-CS-BSA溶液中的完整蛋白没有明显变化，但在1 小时后，游离BSA完全被消化。6 后，在GO-BSA和GO-CS-BSA溶液中检测到完整的蛋白质，而在游离的BSA溶液中未检测到完整的蛋白质。
Proteins have short half-life because of enzymatic cleavage. Here, a new protein nanocarrier made of graphene oxide (GO) + Chitosan (CS) is proposed to successfully prevent proteolysis in protein and simultaneously retain its activity. Bovine serum albumin (BSA) and collagenase were loaded on GO and GO-CS to explore the stability and activity of proteins. SEM, AFM, TEM, DSC, UV-Vis, FT-IR, RBS, Raman, SDS-PAGE and zymography were utilized as characterization techniques. The protecting role of GO and GO-CS against enzymatic cleavage was probed by protease digestion analysis on BSA, where the protease solution was introduced to GO-BSA and GO-CS-BSA at 37 °C for 0.5-1-3-6 hours. Characterizations showed the successful synthesis of few layers of GO and the coverage by CS. According to gelatin zymographic analysis, the loaded collagenase on GO and GO-CS lysed the gelatin and created non-staining bands which confirmed the activity of loaded collagenase. SDS-PAGE analysis revealed no significant change in the intact protein in the GO-BSA and GO-CS-BSA solution after 30-minute and 1-hour exposure to protease; however, free BSA was completely digested after 1 hour. After 6 hours, intact proteins were detected in GO-BSA and GO-CS-BSA solutions, while no intact protein was detected in the free BSA solution.