Identification of quinoline scaffolds against emerging drug target glycerophosphodiester phosphodiesterase from Staphylococcus aureus: an integrated in vitro, in silico, and STD-NMR approach.

Glycerophosphodiester phosphodiesterases (GDPDs) are biocatalysts and highly conserved enzymes in both prokaryotic and eukaryotic organisms. GDPD plays an important role in cellular metabolism by hydrolyzing glycerophosphodiester substrates into smaller but useful molecules, such as glycerol-3-phosphate and its corresponding alcohol. GDPD directly affects the pathogenicity of infectious bacteria, including Staphylococcus aureus, and consequently represents a potential therapeutic drug target. In the current study, the GDPD enzyme from vancomycin-resistant S. aureus (VRSA) was heterologously expressed in BL21(DE3)pLysS and purified. Its catalytic activity was assessed on bis(p-nitrophenyl) phosphate (BpNPP), a commonly used non-physiological substrate for phosphodiesterases. 24 compounds based on pharmacologically important aminoquinolines were evaluated for their inhibitory potential against GDPD. Among these, 15 compounds exhibited significant to moderate inhibitory potential with IC50 values ranging from 79.3 ± 4.8 to 943.5 ± 17.3 µM. Notably, compound 6 exhibited the most potent inhibition (IC50 = 79.3 ± 4.8 µM), as compared to the standard ethylenediaminetetraacetic acid EDTA (IC50 = 468.5 ± 1.09 µM). Circular dichroism (CD) spectroscopy indicates that GDPD has a properly folded secondary structure, which is essential for enzyme activity. CD spectra showed a red shift upon inhibitor addition, from 211 to 237 nm, reflecting alterations in the enzyme's secondary structure. The thermal shift assay was performed to observe the thermal stability and structural integrity of the enzyme in the presence of inhibitors. Saturation transfer difference (STD) NMR spectroscopy provided limited but key insights into the binding interactions of inhibitors with the target enzyme. Binding interactions were further explored using extensive molecular docking studies.