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Abstract
Biofilms are important for cell communication and growth in most bacteria, and are responsible for a number of human clinical infections and diseases. TpbA (PA3885) is a dual specific tyrosine phosphatase (DUSP) that negatively regulates biofilm formation in the opportunistic pathogen Pseudomonas aeruginosa PAO1 by converting extracellular quorum sensing signals into internal gene cascade reactions that result in reduced biofilm formation. We have determined the three-dimensional crystal structure of wild-type TpbA from P. aeruginosa PAO1 in the phosphate-bound state and a TpbA (C132S) mutant with phosphotyrosine. Comparison between the phosphate-bound structure and the previously reported ligand-free TpbA structure reveals the extent of conformational changes that occur upon substrate binding. The largest changes occur in the functional loops that define the substrate binding site, including the PTP, general acid and α4-α5 loops. We further show that TpbA efficiently catalyzes the hydrolysis of two phosphotyrosine peptides derived from the periplasmic domain of TpbB (YfiN, PA1120), with a strong preference for dephosphorylating Tyr48 over Tyr62. This work adds to the small repertoire of DUSP structures in both the ligand-free and ligand-bound states, and provides a starting point for further study of the role of TpbA in biofilm formation.
Highlights
Pseudomonas aeruginosa, a bacterium noted for its metabolic variety, is found in a wide range of biotic and abiotic habitats including water, soil and various organisms
Comparison with the recent solution structure of TpbA in the open ligand-free state reveals the extent of the conformational changes that occur upon ligand binding. This adds to the small repertoire of dual specific tyrosine phosphatase (DUSP) structures that have been determined in both the ligand-free and ligand-bound states, and should provide greater understanding of the relationship between protein dynamics and phosphatase activity and substrate selectivity
TpbA has been shown to act as a negative regulator of cellular c-di-GMP formation, and loss of TpbA leads to increased concentrations of c-di-GMP that result in enhancement of biofilm formation and inhibition of motility
Summary
Pseudomonas aeruginosa, a bacterium noted for its metabolic variety, is found in a wide range of biotic and abiotic habitats including water, soil and various organisms. Structure and Biochemistry of P. aeruginosa TpbA collection and analysis, decision to publish, or preparation of the manuscript
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