Functionalization of Cellulose-Based Hydrogels with Bi-Functional Fusion Proteins Containing Carbohydrate-Binding Modules.

Mariana Barbosa,Duarte Miguel F Prazeres,Hélvio Simões

doi:10.3390/ma14123175

Mariana Barbosa, Duarte Miguel F Prazeres + Show 1 more

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https://doi.org/10.3390/ma14123175

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Abstract

Materials with novel and enhanced functionalities can be obtained by modifying cellulose with a range of biomolecules. This functionalization can deliver tailored cellulose-based materials with enhanced physical and chemical properties and control of biological interactions that match specific applications. One of the foundations for the success of such biomaterials is to efficiently control the capacity to combine relevant biomolecules into cellulose materials in such a way that the desired functionality is attained. In this context, our main goal was to develop bi-functional biomolecular constructs for the precise modification of cellulose hydrogels with bioactive molecules of interest. The main idea was to use biomolecular engineering techniques to generate and purify different recombinant fusions of carbohydrate binding modules (CBMs) with significant biological entities. Specifically, CBM-based fusions were designed to enable the bridging of proteins or oligonucleotides with cellulose hydrogels. The work focused on constructs that combine a family 3 CBM derived from the cellulosomal-scaffolding protein A from Clostridium thermocellum (CBM3) with the following: (i) an N-terminal green fluorescent protein (GFP) domain (GFP-CBM3); (ii) a double Z domain that recognizes IgG antibodies; and (iii) a C-terminal cysteine (CBM3C). The ability of the CBM fusions to bind and/or anchor their counterparts onto the surface of cellulose hydrogels was evaluated with pull-down assays. Capture of GFP-CBM3 by cellulose was first demonstrated qualitatively by fluorescence microscopy. The binding of the fusion proteins, the capture of antibodies (by ZZ-CBM3), and the grafting of an oligonucleotide (to CBM3C) were successfully demonstrated. The bioactive cellulose platform described here enables the precise anchoring of different biomolecules onto cellulose hydrogels and could contribute significatively to the development of advanced medical diagnostic sensors or specialized biomaterials, among others.

Highlights

New functional cellulose-based materials can be obtained through the assembly of bioactive molecules that enhance the existing properties or add novel potentialities [1].Such matrices have the potential to be applied in a great number of different fields, ranging from functional textiles [2], smart packaging [3], and biomanufacturing [4] to biosensing [5]and advanced drug delivery systems [6].Cellulose is the most abundant natural biopolymer on Earth and, one of the most important biomass resources [7]
We demonstrated that biomolecular recognition is a successful approach for the functionalization of cellulose-based hydrogels with a fluorescent protein, a protein domain that recognizes immunoglobulin G (IgG) antibodies, and a C-terminal cysteine to covalently link DNA strands
The results suggest that these CBM3-based fusions were able to efficiently bind cellulose even if it is present in a hydrogel form

Summary

Introduction

New functional cellulose-based materials can be obtained through the assembly of bioactive molecules that enhance the existing properties or add novel potentialities [1].Such matrices have the potential to be applied in a great number of different fields, ranging from functional textiles [2], smart packaging [3], and biomanufacturing [4] to biosensing [5]and advanced drug delivery systems [6].Cellulose is the most abundant natural biopolymer on Earth and, one of the most important biomass resources [7]. New functional cellulose-based materials can be obtained through the assembly of bioactive molecules that enhance the existing properties or add novel potentialities [1]. Such matrices have the potential to be applied in a great number of different fields, ranging from functional textiles [2], smart packaging [3], and biomanufacturing [4] to biosensing [5]. Cellulose is the most abundant natural biopolymer on Earth and, one of the most important biomass resources [7] This biopolymer forms highly crystalline structures with elongated stiff chain packing due to the β(1,4)-linked glycosidic arrangement of glucose repeating units and acts as a structural material in biological systems [7,8]. Cellulose acetate, a well-known derivative of cellulose produced by acetylation of native cellulose, has a much less crystalline structure, and exhibits better solubility in conventional solvents such as acetone [13,14]

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