NMR characterization of weak interactions between RhoGDI2 and fragment screening hits

Liu, Jiuyang; Gao, Jia; Li, Fudong; Ma, Ronghua; Wei, Qingtao; Wang, Aidong; Wu, Jihui; Ruan, Ke

doi:10.1016/j.bbagen.2016.10.003

Cited by 16 publications

(16 citation statements)

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“…During primary screening, we screened the saturation transfer difference (STD), WaterLOGSY, and CPMG spectra using fragment cocktails (10 compounds each at 0.4 m m and 10 μ m OptrA with 50% D 2 O in PBS buffer). The interpretation of the four spectra was performed as previously described . Then, we compared reference 1H spectra for each individual compound and the peak multiplicities from the STD, WaterLOGSY, and CPMG spectra for each fragment cocktail to screen for primary hits.…”

Section: Methodsmentioning

confidence: 99%

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A novel inhibitor of the new antibiotic resistance protein OptrA

et al. 2018

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The antibiotic resistance (ARE) subfamily of ABC (ATP-binding cassette) proteins confers resistance to a variety of clinically important ribosome-targeting antibiotics and plays an important role in infections caused by pathogenic bacteria. However, inhibitors of ARE proteins have rarely been reported. Here, OptrA, a new member of the ARE proteins, was used to study inhibitors of these types of proteins. We first confirmed that destroying the catalytic activity of OptrA could restore the sensitivity of host cells to antibiotics. Then, fragment-based screening, a drug screening method, was used to screen for inhibitors of OptrA. The competitive saturation transfer difference experiments, docking, and molecular dynamics were used to determine the binding sites and mode of interactions between OptrA and fragment screening hits. In this study, we first find a novel and specific inhibitor of OptrA (CP1), which suppressed the ATPase activity of OptrA in vitro by 30%. A hydrogen bond formed between the 8-position phenylcyclic cyano group in CP1 and the amino acid residue Lys-271 allows CP1 to form a stable complex with OptrA protein. These findings provide a theoretical basis for the further optimization of the inhibitor structure to obtain inhibitors with higher efficiencies.

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Section: Methodsmentioning

confidence: 99%

“…FBS is a sensitive biophysical technique used to detect the binding of small molecules to protein targets using a special small molecular library . Despite a low sensitivity, NMR spectroscopy is a robust screening method of high information contents which will help us to screen out molecular drugs with higher specificity …”

Section: Introductionmentioning

confidence: 99%

A novel inhibitor of the new antibiotic resistance protein OptrA

et al. 2018

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“…The 15 N-labeled AtPsb27 was used for the heteronuclear single quantum coherence (HSQC) spectrum analysis followed the procedure used by Liu et al ( 2017 ). Protein was concentrated to 0.2 mM in a buffer containing 20 mM sodium phosphate, 250 mM NaCl, 2.5 mM DTT, 2.5 mM Na 2 EDTA, and 0.002% NaN 3 at pH6.7, 7.0, and 7.5, respectively.…”

Section: Methodsmentioning

confidence: 99%

“…Protein crystallization and structure determination were conducted following a protocol previously described by Liu et al ( 2017 ). In brief, the purified AtPsb27 was dialyzed against a buffer of 20 mM Tris–HCl (pH 8.0) and 200 mM NaCl at 4 °C and then concentrated to 22 mg/ml using an Amicon Centricon YM-10 (Millipore, Billerica, MA, USA).…”

Section: Methodsmentioning

confidence: 99%

Crystal structure of Psb27 from Arabidopsis thaliana determined at a resolution of 1.85 Å

et al. 2017

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Proper biogenesis and maintenance of photosynthetic thylakoid membrane complexes are essential for the photosynthetic light reactions. A thylakoid lumenal protein, Psb27, plays a vital role in assembly or/and maintenance of photosystem II (PSII). In cyanobacteria, it is a small lipoprotein docked to the lumenal side of PSII, and functions in the assembly of the Mn4Ca cluster and in the PSII repair cycle. However, Psb27 from Arabidopsis thaliana is not a lipoprotein, and it is involved in PSII repair and acclimation to fluctuating light stress, suggesting a functional divergence between Arabidopsis Psb27 and cyanobacterial Psb27s. To gain a better understanding of Psb27 from higher plants, we determined the crystal structure of Arabidopsis Psb27 by X-ray crystallography at a resolution of 1.85 Å. The structure of Arabidopsis Psb27 is a four-helix bundle, similar to its orthologues from cyanobacteria. However, there are several structural differences between Arabidopsis Psb27 and cyanobacterial Psb27s concerning the overall molecular shape, the N- and C-terminal structures, and the surface charge. These differences suggest that Psb27 from higher plants and cyanobacteria may function differently.Electronic supplementary materialThe online version of this article (doi:10.1007/s11120-017-0450-3) contains supplementary material, which is available to authorized users.

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“…These weak binders can be readily identified using a nuclear magnetic resonance (NMR) fragment-based approach, thereby eliminating the distracting false positives in virtual screening. Using our NMR fragment-based approach [16][17][18] , we characterized weak target-ligand interactions using ligand-observed spectra, e.g., saturation transfer difference (STD) and WaterLOGSY. Three hits for the SARS-CoV-2 M pro were identified ( Figure 1a): niacin, hit 1, and hit 2 (Figure 1b).…”

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Repurposing low–molecular-weight drugs against the main protease of severe acute respiratory syndrome coronavirus 2

Gao

Zhang

Liu

et al. 2020

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The coronavirus disease pandemic caused by infection with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has affected the global healthcare system. Drug repurposing is a feasible method for emergency treatment. As low-molecular-weight drugs have high potential to completely match interactions with essential SARS-CoV-2 targets, we propose a strategy to identify such drugs using the fragment-based approach. Herein, using ligand-and protein-observed fragment screening approaches, we identified niacin and hit 1 binding to the catalytic pocket of the main protease of the SARS-CoV-2 (M pro ), thereby modestly inhibiting the enzymatic activity of M pro . Chemical shift perturbations induced by niacin and hit 1 indicate a partial overlap of their binding sites, i.e., the catalytic pocket of M pro may accommodate derivatives with large molecular sizes. Therefore, we searched for drugs containing niacin or hit 1 pharmacophores and identified carmofur, bendamustine, triclabendazole, and emedastine; these drugs are highly capable of inhibiting protease activity. Our study demonstrates that the fragment-based approach is a feasible strategy for identifying low-molecular-weight drugs against the SARS-CoV-2 and other potential targets lacking specific drugs.

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NMR characterization of weak interactions between RhoGDI2 and fragment screening hits

Cited by 16 publications

References 50 publications

A novel inhibitor of the new antibiotic resistance protein OptrA

A novel inhibitor of the new antibiotic resistance protein OptrA

Crystal structure of Psb27 from Arabidopsis thaliana determined at a resolution of 1.85 Å

Repurposing low–molecular-weight drugs against the main protease of severe acute respiratory syndrome coronavirus 2

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