Your browser doesn't support javascript.
loading
Anionic amino acids support hydrolysis of poly-ß-(1,6)-N-acetylglucosamine exopolysaccharides by the biofilm dispersing glycosidase Dispersin B.
Breslawec, Alexandra P; Wang, Shaochi; Li, Crystal; Poulin, Myles B.
Affiliation
  • Breslawec AP; Department of Chemistry and Biochemistry, University of Maryland, College Park, Maryland, USA.
  • Wang S; Department of Chemistry and Biochemistry, University of Maryland, College Park, Maryland, USA.
  • Li C; Department of Chemistry and Biochemistry, University of Maryland, College Park, Maryland, USA.
  • Poulin MB; Department of Chemistry and Biochemistry, University of Maryland, College Park, Maryland, USA. Electronic address: mpoulin@umd.edu.
J Biol Chem ; 296: 100203, 2021.
Article in En | MEDLINE | ID: mdl-33334876
The exopolysaccharide poly-ß-(1→6)-N-acetylglucosamine (PNAG) is a major structural determinant of bacterial biofilms responsible for persistent and nosocomial infections. The enzymatic dispersal of biofilms by PNAG-hydrolyzing glycosidase enzymes, such as Dispersin B (DspB), is a possible approach to treat biofilm-dependent bacterial infections. The cationic charge resulting from partial de-N-acetylation of native PNAG is critical for PNAG-dependent biofilm formation. We recently demonstrated that DspB has increased catalytic activity on de-N-acetylated PNAG oligosaccharides, but the molecular basis for this increased activity is not known. Here, we analyze the role of anionic amino acids surrounding the catalytic pocket of DspB in PNAG substrate recognition and hydrolysis using a combination of site-directed mutagenesis, activity measurements using synthetic PNAG oligosaccharide analogs, and in vitro biofilm dispersal assays. The results of these studies support a model in which bound PNAG is weakly associated with a shallow anionic groove on the DspB protein surface with recognition driven by interactions with the -1 GlcNAc residue in the catalytic pocket. An increased rate of hydrolysis for cationic PNAG was driven, in part, by interaction with D147 on the anionic surface. Moreover, we identified that a DspB mutant with improved hydrolysis of fully acetylated PNAG oligosaccharides correlates with improved in vitro dispersal of PNAG-dependent Staphylococcus epidermidis biofilms. These results provide insight into the mechanism of substrate recognition by DspB and suggest a method to improve DspB biofilm dispersal activity by mutation of the amino acids within the anionic binding surface.
Subject(s)
Key words

Full text: 1 Collection: 01-internacional Database: MEDLINE Main subject: Bacterial Proteins / Aggregatibacter actinomycetemcomitans / Beta-Glucans / Amino Acids / Glycoside Hydrolases Language: En Journal: J Biol Chem Year: 2021 Document type: Article Affiliation country: Estados Unidos Country of publication: Estados Unidos

Full text: 1 Collection: 01-internacional Database: MEDLINE Main subject: Bacterial Proteins / Aggregatibacter actinomycetemcomitans / Beta-Glucans / Amino Acids / Glycoside Hydrolases Language: En Journal: J Biol Chem Year: 2021 Document type: Article Affiliation country: Estados Unidos Country of publication: Estados Unidos