Functionalized Graphene Oxide with Chitosan for Protein Nanocarriers to Protect against Enzymatic Cleavage and Retain Collagenase Activity

Abstract

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.

Fabrication of a Biocompatible Liquid Crystal Graphene Oxide–Gold Nanorods Electro‐ and Photoactive Interface for Cell Stimulation

“…The AuNR showed peaks at 3451, 2868, 1742, 1654, 1460 and 1103 cm −1 corresponding to NH/OH, CH, CO, amide I, amide II, CO groups present in the protein coating of the gold nanorods, respectively . The success of the EDC/NHS conjugation was confirmed firstly by the presence of characteristic amide‐carbonyl (NHCO) stretching (amide I) at 1621 cm −1 in LCGO–AuNR . Second, the disappearance of carboxylic acid stretch (CO) in LCGO–AuNR at 1710 cm −1 indicated that the COOH group of LCGO were consumed in the conjugation to form the amide linkage (Figure b arrow) .…”

“…As shown in Figure b, the FTIR spectrum of LCGO presented peaks at 3390, 1710, 1630, 1220, and 1050 cm −1 , which were attributed to the OH bond stretch, CO in carboxylic acid, CC, COC, and CO stretch groups, respectively . The AuNR showed peaks at 3451, 2868, 1742, 1654, 1460 and 1103 cm −1 corresponding to NH/OH, CH, CO, amide I, amide II, CO groups present in the protein coating of the gold nanorods, respectively . The success of the EDC/NHS conjugation was confirmed firstly by the presence of characteristic amide‐carbonyl (NHCO) stretching (amide I) at 1621 cm −1 in LCGO–AuNR .…”

“…The success of the EDC/NHS conjugation was confirmed firstly by the presence of characteristic amide‐carbonyl (NHCO) stretching (amide I) at 1621 cm −1 in LCGO–AuNR . Second, the disappearance of carboxylic acid stretch (CO) in LCGO–AuNR at 1710 cm −1 indicated that the COOH group of LCGO were consumed in the conjugation to form the amide linkage (Figure b arrow) . The appearance of CH stretch peaks at 2800 cm −1 in LCGO–AuNR, resulting from the CH 2 groups of the gold nanorods' protein‐coating present in the nanocomposite, further confirmed the conjugation.…”

“…FTIR was performed to confirm the EDC/NHS conjugation of LCGO to AuNR, illustrated in Figure a. As shown in Figure b, the FTIR spectrum of LCGO presented peaks at 3390, 1710, 1630, 1220, and 1050 cm −1 , which were attributed to the OH bond stretch, CO in carboxylic acid, CC, COC, and CO stretch groups, respectively . The AuNR showed peaks at 3451, 2868, 1742, 1654, 1460 and 1103 cm −1 corresponding to NH/OH, CH, CO, amide I, amide II, CO groups present in the protein coating of the gold nanorods, respectively .…”

Electrochemical sensing of glucose by chitosan modified graphene oxide

“…ascorbic acid, is quite limited. Several examples reported the added value of using GO and Ch blends: Ch possesses a high number of amine residues that can be exploited for covalent grafting of GO [35][36][37][38][39][40] through amidation reaction with the carboxylic moieties or epoxide ring opening [41]. While amidation requires activation of carboxyls, the epoxide ring opening by primary amines can occur more easily, without activation, but it strongly depends on the amine structure, reaction time, pH and solvent [42]; for example, reaction of GO with polyethyleneimide occurred in water:EtOH 1:1 mixture under reflux for 24 h [43], reaction with TEG diamine occurred in DMF after five days [36], reaction with EDTA occurred in EtOH under sonication for 24 h at room temperature [44].…”