Browsing Department of Drug design and optimization ([HIPS]DDOP) by Subjects
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Protein-observed 19F NMR of LecA from Pseudomonas aeruginosa.The carbohydrate-binding protein LecA (PA-IL) from Pseudomonas aeruginosa plays an important role in the formation of biofilms in chronic infections. Development of inhibitors to disrupt LecA-mediated biofilms is desired, but limited to carbohydrate-based ligands. Moreover, discovery of drug-like ligands for LecA is challenging due to its weak affinities. Therefore, we established a protein-observed 19F (PrOF) NMR to probe ligand binding to LecA. LecA was labeled with 5 - fluoroindole to incorporate 5 - fluorotryptophanes and the resonances were assigned by site-directed mutagenesis. This incorporation did not disrupt LecA preference for natural ligands, Ca2+ and d - galactose. Following NMR resonance perturbation of W42, which is located in the carbohydrate-binding region of LecA, allowed to monitor binding of low affinity ligands such as N - acetyl d - galactosamine (d - GalNAc, Kd = 780 ± 97 μM). Moreover, PrOF NMR titration with glycomimetic of LecA p-nitrophenyl β-d-galactoside (pNPGal, Kd = 54 ± 6 μM) demonstrated a six-fold improved binding of d - Gal proving this approach to be valuable for ligand design in future drug discovery campaigns that aim to generate inhibitors of LecA.
Search for the Active Ingredients from a 2-Aminothiazole DMSO Stock Solution with Antimalarial Activity.Chemical decomposition of DMSO stock solutions is a common incident that can mislead biological screening campaigns. Here, we share our case study of 2-aminothiazole 1, originating from an antimalarial class that undergoes chemical decomposition in DMSO at room temperature. As previously measured biological activities observed against Plasmodium falciparum NF54 and for the target enzyme PfIspE were not reproducible for a fresh batch, we tackled the challenge to understand where the activity originated from. Solvent- and temperature-dependent studies using HRMS and NMR spectroscopy to monitor the decomposition led to the isolation and in vitro evaluation of several fractions against PfIspE. After four days of decomposition, we successfully isolated the oxygenated and dimerised compounds using SFC purification and correlated the observed activities to them. Due to the unstable nature of the two isolates, it is likely that they undergo further decomposition contributing to the overall instability of the compound.