Structural Polymorphism of Chitin and Chitosan in Fungal Cell Walls From Solid-State NMR and Principal Component Analysis.

Liyanage D Fernando,Malitha C Dickwella Widanage,Liqun Zhang,Jean-Paul Latgé,Andrew S Lipton,Jackson Penfield,Tuo Wang,Nancy Washton,Ping Wang

doi:10.3389/fmolb.2021.727053

Abstract

Chitin is a major carbohydrate component of the fungal cell wall and a promising target for novel antifungal agents. However, it is technically challenging to characterize the structure of this polymer in native cell walls. Here, we recorded and compared 13C chemical shifts of chitin using isotopically enriched cells of six Aspergillus, Rhizopus, and Candida strains, with data interpretation assisted by principal component analysis (PCA) and linear discriminant analysis (LDA) methods. The structure of chitin is found to be intrinsically heterogeneous, with peak multiplicity detected in each sample and distinct fingerprints observed across fungal species. Fungal chitin exhibits partial similarity to the model structures of α- and γ-allomorphs; therefore, chitin structure is not significantly affected by interactions with other cell wall components. Addition of antifungal drugs and salts did not significantly perturb the chemical shifts, revealing the structural resistance of chitin to external stress. In addition, the structure of the deacetylated form, chitosan, was found to resemble a relaxed two-fold helix conformation. This study provides high-resolution information on the structure of chitin and chitosan in their cellular contexts. The method is applicable to the analysis of other complex carbohydrates and polymer composites.

Highlights

Chitin is the second-most abundant biopolymer in nature, only behind cellulose
The nitrogenated sugars in the intracellular content have already been filtered out using cross polarization (CP)-based methods, which remove the signals of mobile sugars but selectively highlight the stiff molecules in the cell wall
High-resolution 2D 13C-13C and 15N-13C correlation spectra collected on freshly prepared A. fumigatus mycelia resolved the signals of six major types of chitins, together with two forms with some carbon sites being ambiguously assigned (Figure 2A; Supplementary Figure S2)

Summary

Introduction

Chitin is the second-most abundant biopolymer in nature, only behind cellulose. Widely distributed in different organisms, chitin is often found as a supportive and protective component of the body armor (namely the exoskeleton) in arthropods and the cell walls of fungi and some algal species (Pillai et al, 2009; Rinaudo, 2006). The structures of chitin and its largely deacetylated form called chitosan have similarity to the organization of cellulose (Heux et al, 2000; Jarvis, 2003; Okuyama et al, 2000; Rinaudo, 2006; Saito et al, 1987). All these three polysaccharides are linear polymers of β-1,4-linked glucoses or their amide derivatives. N-acetylglucosamine (GlcNAc) unit in chitin and the glucosamine (GlcN) residue in chitosan (Figure 1A). Especially the latter, have drawn tremendous attention due to their promising applications as polymer scaffolds for tissue engineering, wound dressing, drug delivery, and pharmaceuticals (Jayakumar et al, 2010)

Methods

Results

Conclusion