Genetic analysis of cellulose crystallization in Gluconacetobacter xylinus

In nature, polysaccharides, such as cellulose, are important biopolymers involved in numerous biological functions including prevention of cellular desiccation, energy storage, osmoregulation, and cell wall formation. Apart from having a relevant biological role, cellulose also has a great economic importance. For instance, cellulose provides the raw material for paper and textile production, and it is predicted to become a key precursor for glucose-derived ethanol. The mechanisms underlying cellulose biosynthesis are not completely understood. Elucidation of these mechanisms are essential for efficient cellulose production and industrial applications. In an effort to gain a better understanding of the cellulose biosynthesis process and its regulation, the crystallization of cellulose was investigated in Gluconacetobacter xylinus mutants resistant to a novel inhibitor of cellulose I formation known as pellicin. Through the use of forward genetics and site-directed mutagenesis, the mutations A449T and A449V in the G. xylinus BcsA protein were found to be important to confer high levels of pellicin resistance. Phenotypic analysis of the BcsA A449T and BcsA A449V mutants did not reveal other apparent phenotypic alterations when compared to the wild type. This suggests that these mutations are exclusively involved in cellulose crystallization. The localization of the A449 amino acid residue in the BcsA protein and structural analysis of both the native and the mutant BcsA proteins, by means of 3D modelling, suggest that pellicin may inhibit cellulose crystallization either by affecting the translocation process of the glucan within the BcsA protein or by allosterically altering protein-protein interactions