Identification of two dihydrodipicolinate synthase isoforms from Pseudomonas aeruginosa that differ in allosteric...

Area of Science:

• Microbiology
• Biochemistry
• Drug Discovery

Background:

• Pseudomonas aeruginosa is a major cause of hospital-acquired infections, with rising antibiotic resistance necessitating novel therapeutic targets.
• The diaminopimelate (DAP) pathway is essential for bacterial cell wall and protein biosynthesis, making its enzymes attractive targets for new antibiotics.
• Dihydrodipicolinate synthase (DHDPS) catalyzes the rate-limiting step in the DAP pathway and is typically encoded by a single gene (dapA).

Purpose of the Study:

• To investigate the presence and characteristics of DHDPS enzymes in P. aeruginosa.
• To understand the functional and regulatory differences between P. aeruginosa DHDPS isoforms.
• To identify potential new targets for combating P. aeruginosa infections.

Main Methods:

• Bioinformatic analysis to identify DHDPS genes.
• Enzyme activity assays to determine catalytic rates (kcat).
• Allosteric inhibition studies using lysine to determine IC50 values.
• Structural analysis to identify key amino acid differences.

Main Results:

• P. aeruginosa encodes two functional DHDPS enzymes, PaDHDPS1 and PaDHDPS2.
• Both enzymes exhibit similar catalytic activities.
• PaDHDPS2 is allosterically inhibited by lysine, unlike PaDHDPS1, due to a single amino acid difference in the allosteric binding pocket.
• This is the first report of a bacterium with multiple, differentially regulated DHDPS isoforms.

Conclusions:

• The presence of two DHDPS isoforms with distinct regulatory mechanisms in P. aeruginosa may enhance metabolic flux through the DAP pathway.
• These findings provide a basis for developing novel antibiotics targeting the DAP pathway in P. aeruginosa.
• Understanding the differential regulation of PaDHDPS1 and PaDHDPS2 is critical for designing effective inhibitors.