Model of PNAG biosynthetic machinery
Poly-β-1,6-N-acetyl-D-glucosamine (PNAG) is a key virulence factor required for biofilm formation by numerous pathogenic Gram-positive and Gram-negative bacteria. The polymer has been implicated in septicemia, prosthetic joint infection, infective endocarditis, urinary tract infections, whopping cough, and neonatal bacteremia. Partial deacetylation of PNAG (dPNAG) is required for the structural development and integrity of the biofilm. De-N-acetylation of PNAG occurs in the periplasm in Gram-negative species, and deletion of the deacetylase gene has different effects depending on the organism. For example, E. coli strains lacking pgaB cannot export PNAG from the cell, while Bordetella bronchiseptica bpsB deletion strains have a biofilm-defective phenotype and significantly altered biofilm architecture. In contrast, in Gram-positive species deacetylation occurs extracellularly, and deletion of the deacetylase icaB results in shedding of the polymer from the cell surface. As a result strains that lack the deacetylase have highly attenuated virulence.
Given the importance of this chemical modification for biofilm formation, we have focused our efforts on understanding the structure and function of the deacetylase enzymes, PgaB, BpsB and IcaB from E. coli, B. bronchiseptica, and S. epidermidis, respectively. Structural analyses have shed light on unique features of each enzyme and have suggested a role for C-terminal domain of PgaB/BpsB in both efficient deacetylation and polymer export.
Current projects
Ongoing projects are focused on:
- Exploring the role of the C-terminal domain of PgaB/BpsB in polymer biosynthesis.
- Understanding how the polymer is synthesized and transported across the cytoplasmic membrane.
- Determining how the proteins PgaA/BpsA and PgaB/BpsB interact to facilitate export of the polymer from the cell.
- Developing small molecule modulators of deacetylase activity.
Selected Publications
- The protein BpsB is a poly-β-1,6-N-acetyl-D-glucosamine deacetylase required for biofilm formation in Bordetella bronchiseptica. Little DJ, Milek S, Bamford NC, Ganguly T, DiFrancesco BR, Nitz M, Deora R, Howell PL. J Biol Chem. 2015 Sep 11;290(37):22827-40. doi: 10.1074/jbc.M115.672469.
- Structural basis for the De-N-acetylation of Poly-β-1,6-N-acetyl-D-glucosamine in Gram-positive bacteria. Little DJ, Bamford NC, Pokrovskaya V, Robinson H, Nitz M, Howell PL. J Biol Chem. 2014 Dec 26;289(52):35907-17. doi: 10.1074/jbc.M114.611400.
- Modification and periplasmic translocation of the biofilm exopolysaccharide poly-β-1,6-N-acetyl-D-glucosamine. Little DJ, Li G, Ing C, DiFrancesco BR, Bamford NC, Robinson H, Nitz M, Pomès R, Howell PL. Proc Natl Acad Sci U S A. 2014 Jul 29;111(30):11013-8. doi: 10.1073/pnas.1406388111.