Submitted to: Cellulose
Publication Type: Peer reviewed journal
Publication Acceptance Date: 12/6/2011
Publication Date: 4/1/2012
Citation: Edwards, J.V., Prevost, N.T., Condon, B.D., French, A.D., Wu, Q. 2012. Immobilization of lysozyme-cellulose amide-linked conjugates on cellulose I and II cotton nanocrystalline preparations. Cellulose. 19(2):495-506. Interpretive Summary: The creation of new markets for value-added, protective cotton textiles is currently an agriculture issue. Work in Southern Regional Research Center’s Cotton Chemistry Utilization Unit is addressing the application of bio-active molecules to cotton textiles as a model to explore the creation of new value-added cotton products with highly selective properties and environmentally acceptable finishes. As an example of a protein’s biological activity on cotton, enzyme conjugates of cellulose on cotton performance fabrics have been studied for their antimicrobial activity. In this paper we explore the attachment of lysozyme to cotton nanocrystals, which were prepared and characterized as two types of molecular structures – cellulose I and II. The loading of enzyme on nano-sized matrices provides an ideal vehicle for presenting highly concentrated and stable enzyme activity. The immobilization chemistry in this study gave extraordinary incorporation of lysozyme activity. These types of immobilized enzyme formulations are being explored for applications in numerous areas including antimicrobial, bio-remediation, bio-fouling, and bio-sensing.
Technical Abstract: Lysozyme was attached through an amide linkage between protein aspartate and glutamate residues to amino-glycine-cellulose (AGC), which was prepared by esterification of glycine to preparations of cotton nanocrystals (CNC). The nanocrystalline preparations were produced through acid hydrolysis and mechanical breakage of the cotton fibers from a scoured and bleached cotton fabric and a scoured and bleached, mercerized fabric, which was shown to produce cellulose I (NCI) and cellulose II (NCII) crystals respectively. A carbodiimide-activation coupling reaction was used to create the lysozyme-amino-glycine-cellulose conjugates using both NCI and NCII in a polar solvent, and giving yields of covalently linked lysozyme at 630mg/gram of cotton nanocrystal. The incorporation of lysozyme conjugated to the NCI and NCII preparations gave extraordinarily high activity (1500 U/ mg cotton) when assessed using a fluorescence tag assay to measure antimicrobial activity against Micrococcus lysodeikticus. Scanning electron micrographs demonstrated an aggregation of nanoparticles corresponding to lysozyme bound on the surface of larger cotton nanocrystalline sheets. The approach of producing high enzyme activity on cotton nanocrystals is discussed in the context of selectively presenting robust hydrolase activity on nanocrystalline surfaces.