Methionine aminopeptidase (MetAP) is a class of ubiquitous enzymes essential for the survival of numerous bacterial species. These enzymes are responsible for the cleavage of N-terminal formyl-methionine initiators from nascent proteins to initiate post-translational modifications that are often essential to proper protein function. Thus, inhibition of MetAP activity has been implicated as a novel antibacterial target. We tested this idea in the present study by targeting the MetAP enzyme in the obligate intracellular pathogen Rickettsia prowazekii. We first identified potent RpMetAP inhibitory species by employing an in vitro enzymatic activity assay. The molecular docking program AutoDock was then utilized to compare published crystal structures of inhibited MetAP species to docked poses of RpMetAP. Based on these in silico and in vitro screens, a subset of 23 compounds was tested for inhibition of R. prowazekii growth in a pulmonary vascular endothelial cell (EC) culture infection model system. All compounds were tested over concentration ranges that were determined to be non-toxic to the ECs and 10 of the 23 compounds displayed substantial inhibition of R. prowazekii growth. These data highlight the therapeutic potential for inhibiting RpMetAP as a novel antimicrobial strategy and set the stage for future studies in pre-clinical animal models of infection.
Methionine aminopeptidase (MetAP) is a dinuclear metalloprotease responsible for the cleavage of methionine initiator residues from nascent proteins. MetAP activity is necessary for bacterial proliferation and is therefore a projected novel antibacterial target. A compound library consisting of 294 members containing metal-binding functional groups was screened against Rickettsia prowazekii MetAP to determine potential inhibitory motifs. The compounds were first screened against the target at a concentration of 10 µM and potential hits were determined to be those exhibiting greater than 50% inhibition of enzymatic activity. These hit compounds were then rescreened against the target in 8-point dose-response curves and 11 compounds were found to inhibit enzymatic activity with IC values of less than 10 µM. Finally, compounds (1-5) were docked against RpMetAP with AutoDock to determine potential binding mechanisms and the results were compared with crystal structures deposited within the PDB.
AP-2 gamma (AP-2γ) is a transcription factor that plays pivotal roles in breast cancer biology. To search for small molecule inhibitors of AP-2γ, we performed a high-throughput fluorescence anisotropy screen and identified a polyoxometalate compound with Wells-Dawson structure K[PMoO] (Dawson-POM) that blocks the DNA-binding activity of AP-2γ. We showed that this blocking activity is due to the direct binding of Dawson-POM to AP-2γ. We also provided evidence to show that Dawson-POM decreases AP-2γ-dependent transcription similar to silencing the gene. Finally, we demonstrated that Dawson-POM contains anti-proliferative and pro-apoptotic effects in breast cancer cells. In summary, we identified the first small molecule inhibitor of AP-2γ and showed Dawson-POM-mediated inhibition of AP-2γ as a potential avenue for cancer therapy.
Aim The association between sugar‐sweetened beverages and metabolic disorders has been well studied. However, it has not been determined whether fasting serum fructose is associated with metabolic dysfunction‐associated fatty liver disease (MAFLD). Methods Participants were enrolled from 2011 to 2012 in Shanghai. Fasting serum fructose concentration was measured with a validated liquid chromatography–tandem mass spectrometry method. Results A total of 954 participants without diabetes were included. They were followed for an average of 3.5 years. A total of 320 (33.5%) participants had MAFLD at baseline. With the increase in fasting serum fructose level by quartile, the MAFLD prevalence was increased by 27.0%, 25.0%, 37.4%, and 44.5%, respectively (p < 0.001). Each SD increase in fasting serum fructose level was associated with a 60% increased risk of MAFLD (odds ratio 1.60; 95% confidence interval [CI], 1.36–1.88; p < 0.001). Fasting serum fructose levels were more closely associated with four components of MAFLD (hepatic steatosis, prediabetes, insulin resistance, and low high‐density lipoprotein). We built a diagnostic model named the fructose fat index (FFI). The area under the receiver operating characteristic curve of the FFI was 0.879 (95% CI, 0.850–0.908) in the derivation cohort and 0.827 (95% CI, 0.776–0.878) in the validation cohort. Subsequent prospective studies found that the incidence risk of MAFLD was 2.26 times higher in the high‐fructose group than in the low‐fructose group among female participants (95% CI, 1.46–3.49; p < 0.001). Conclusion Fasting serum fructose concentration, which mostly reflects endogenous fructose, was associated with a higher risk of MAFLD. The FFI derived from fasting serum fructose could be used to predict MAFLD.
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