Ugd Is Involved in the Synthesis of Glycans of Glycoprotein and LPS and Is Important for Cellulose Degradation in Cytophaga hutchinsonii

<i>Cytophaga hutchinsonii</i>, a member of the phylum <i>Bacteroidetes</i>, can rapidly degrade crystalline cellulose through direct cell-to-substrate contact. Most of its cellulases are secreted by the Type IX secretion system (T9SS) and anchored to the cell surface. Our previous study proved that the C-terminal domain (CTD) of the T9SS substrate cellulase Cel9A is glycosylated in <i>C. hutchinsonii</i>. However, its glycosylation mechanism has remained elusive. In this study, we found that <i>chu_3394</i>, which encodes UDP-glucose 6-dehydrogenase (Ugd), was important for the glycosylation of large amounts of periplasmic and outer membrane proteins in <i>C. hutchinsonii</i>. The contents of mannose, glucose, galactose, and xylose were detected to be reduced in the glycoproteins of the ∆<i>ugd</i> mutant compared to that of wild-type. They might be essential monosaccharides that contribute to the structure and function of glycans attached to proteins in <i>C. hutchinsonii</i>. The depletion of mannose, glucose, galactose, and xylose indicates a decrease in glycosylation modifications in the ∆<i>ugd</i> mutant strain. Then, we found that the deletion of <i>ugd</i> resulted in weakened glycosylation modification of the recombinant green fluorescent protein-tagged CTD of Cel9A. Additionally, the outer-membrane localization of Cel9A was affected in the mutant. Besides this, Ugd was also important for the synthesis of O-antigen of lipopolysaccharide (LPS). Thus, Ugd was involved in the synthesis of glycans in both glycoproteins and LPS in <i>C. hutchinsonii</i>. Moreover, the deletion of <i>ugd</i> affected the cellulose degradation, cell motility, and stress resistance of <i>C. hutchinsonii</i>.