PelA Deacetylase Activity Is Required for Pel Polysaccharide Synthesis in Pseudomonas aeruginosa Academic Article uri icon

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abstract

  • ABSTRACT The Pel polysaccharide serves as an intercellular adhesin for the formation and maintenance of biofilms in the opportunistic pathogen Pseudomonas aeruginosa . Pel biosynthesis requires the products of a seven-gene operon, pelA-pelG , all of which are necessary for Pel-dependent biofilm formation and Pel-related phenotypes. One of the genes, pelA , encodes a protein with a predicted polysaccharide deacetylase domain. In this work, the role of the putative deacetylase domain in Pel production was examined. We first established that purified recombinant PelA hydrolyzed the pseudosubstrate p -nitrophenyl acetate in vitro , and site-specific mutations of predicted deacetylase active-site residues reduced activity greater than 10-fold. Additionally, these mutants were deficient in Pel-dependent biofilm formation and wrinkly colony morphology in vivo . Subcellular fractionation experiments demonstrate that PelA localizes to both the membrane and periplasmic fractions. Finally, antiserum against the Pel polysaccharide was generated, and PelA deacetylase mutants do not produce Pel-reactive material. Taken together, these results suggest that the deacetylase activity of PelA is important for the production of the Pel polysaccharide.

authors

  • Colvin, Kelly M
  • Alnabelseya, Noor
  • Baker, Perrin
  • Whitney, John
  • Howell, P Lynne
  • Parsek, Matthew R

publication date

  • May 15, 2013