Structural and Functional Features of Protein–Polysaccharide Complexes Based on Cysteine Proteases and Hydrophilically Modified Chitosan

Objective: Preparation of protein–polysaccharide complexes allows to modulate practically significant characteristics of both components, thereby increasing not only the practical potential of the resulting structures, but also contributing to our understanding of the features of their interactions. Thus, the aim of this work was to study the structural and functional features of the complexes of cysteine proteases (bromelain, papain and ficin) with a graft copolymer of chitosan with poly(N,N-dimethylaminoethyl methacrylate) (CTS–DMAEMA), and also assess the effect of complexation processes on the protease activity and stability of the enzymes in question. Methods: The CTS–DMAEMA graft copolymer was obtained by solution radical polymerization in 2% acetic acid. Protein–polysaccharide complexes were received in glycine buffer. The protein content in the obtained complexes was determined by the modified Lowry method and proteolytic activity of enzymes as part of complexes was assessed using azocasein as a substrate. Molecular docking was used to identify the types of bonds and interactions within protein–polysaccharide complexes. Results and Discussion: Complexes of cysteine proteases (bromelain, papain, and ficin) with the graft copolymer of chitosan with poly(N,N-dimethylaminoethyl methacrylate) were synthesized. It was found that the complexes have a lower protease activity than the native enzymes, because as a result of complex formation, the active site of cysteine proteases is shielded from the substrate, which complicates the catalysis process. The complexes are more stable compared to the native enzymes and retain up to 26% of protease activity after 21 days of incubation at 37°C in Tris-HCl buffer (pH 7.5). Conclusions: It was found that the proteolytic activity of the complexes is lower compared to their native forms of enzymes. The results of molecular docking showed that the modified chitosan is located in the catalytic pocket of cysteine proteases globules. The synthesized complexes have increased stability on storage under physiological conditions, which makes them promising candidates for the development of wound healing drugs.