Hospital-acquired infections by biofilm forming bacteria continues to be a major health challenge costing an estimated additional $11 billion in health care costs in the USA annually. Bacterial biofilms are sessile surface attached bacteria encased in an extracellular polymeric substance (EPS) consisting of exopolysaccharides, proteins and extracellular DNA that serves to protect the cells and facilitates both cell–cell and cell–surface attachment. Enzymatic breakdown of EPS components has generated interest as a promising strategy to disrupt biofilms. Exopolysaccharides consisting of partially de-N-acetylated β-(1→6)-poly-N-acetylglucosamine(dPNAG) are a common component of the biofilm EPS of both gram-positive and gram-negative human pathogens, including Staphylococcus epidermidis and Staphylococcus aureus. There are only two enzymes that specifically hydrolyze dPNAG reported to date, and relatively little is known about the specific interactions required for substrate recognition and specificity in these enzymes. In this study, we use a combination of chemically defined synthetic dPNAG analogs and targeted site-directed mutagenesis studies to probe the mechanism of substrate recognition by Dispersin B (DspB), a PNAG specific exo-glycosidase. We identified negatively charged patches that play a role in substrate recognition in DspB both in vitro and in biofilm dispersal. These studies provide new insight into the recognition mechanism of deacetylated exopolysaccharides by DspB and their degradation that can aid in the development of more efficient dPNAG hydrolase enzymes.

Chemical and biochemical tools to probe the function of exopolysaccharides in bacterial biofilm formation

ACS Spring 2022

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