Table 2. Peptidoglycan (PGN) quantification through the titration of meso-diaminopimelic acid (mDAP).
Strain | mDAP amounta | Relative PGN amount with regards to PAO1 |
---|---|---|
PAO1 | 215.6 ± 17.5 | 1 |
PAΔDDh2Dh3 | 149.1 ± 9.1* | 0.69* |
PAΔDDh2Dh3 + pUCPAD | 227.6 ± 15.4 | 1.05 |
PAΔDDh2 | 208.9 ± 4.2 | 0.97 |
PAΔDDh3 | 196.5 ± 5.0 | 0.91 |
PAΔDh2Dh3 | 220.3 ± 10.1 | 1.02 |
PAΔDDh2Dh3ΔC | 193.1 ± 13.9 | 0.89 |
PAΔAG | 219.5 ± 16.1 | 1.02 |
PAΔAG + pUCPCAC | 157.5 ± 22.1* | 0.73* |
PAΔAG + pUCPC24 | 191.3 ± 12.0 | 0.89 |
PAΔnZ | 211.5 ± 14.9 | 0.98 |
aThe peptidoglycan from selected strains was quantified by means of the titration of mDAP concentration, following the protocols described in Materials & Methods section. Bacterial cultures previously adjusted to ensure the same number of cells on each.The concentration of muropeptides was calculated using a mDAP standard curve, and the given value is the amount of mDAP (μg) purified from a 250 mL exponential culture adjusted to DO600 = 0.8. The results represent the mean ± SD from three different quantifications per strain.
*Statistically significant: P<0.001 in the One-way ANOVA with post hoc Tukey’s multiple comparison test (with regards to PAO1 strain). PAΔDDh2Dh3: knockout mutant on ampD, ampDh2 and ampDh3 genes; PAΔDDh2Dh3ΔC: knockout mutant on ampD, ampDh2, ampDh3 and ampC genes PAΔnZ: knockout mutant on nagZ. PAΔAG: knockout mutant on ampG. pUCPAD: pUC18-based Escherichia-Pseudomonas shuttle vector containing PAO1 AmpD gene. pUCPAC: pUC18-based Escherichia-Pseudomonas shuttle vector containing PAO1 AmpC gene. pUCP24: empty pUC18-based Escherichia-Pseudomonas shuttle vector.