Flavonoids are plant bioactives that are recognized as hormone-like polyphenols because of their similarity to the endogenous sex steroids 17β-estradiol and testosterone, and to their estrogen- and androgen-like activity. Most efforts to verify flavonoid binding to nuclear receptors (NRs) and explain their action have been focused on ERα, while less attention has been paid to other nuclear and non-nuclear membrane androgen and estrogen receptors. Here, we investigate six flavonoids (apigenin, genistein, luteolin, naringenin, quercetin, and resveratrol) that are widely present in fruits and vegetables, and often used as replacement therapy in menopause. We performed comparative computational docking simulations to predict their capability of binding nuclear receptors ERα, ERβ, ERRβ, ERRγ, androgen receptor (AR), and its variant AR^T877A and membrane receptors for androgens, i.e., ZIP9, GPRC6A, OXER1, TRPM8, and estrogens, i.e., G Protein-Coupled Estrogen Receptor (GPER). In agreement with data reported in literature, our results suggest that these flavonoids show a relevant degree of complementarity with both estrogen and androgen NR binding sites, likely triggering genomic-mediated effects. It is noteworthy that reliable protein–ligand complexes and estimated interaction energies were also obtained for some suggested estrogen and androgen membrane receptors, indicating that flavonoids could also exert non-genomic actions. Further investigations are needed to clarify flavonoid multiple genomic and non-genomic effects. Caution in their administration could be necessary, until the safe assumption of these natural molecules that are largely present in food is assured.

Significance: During infection, the human pathogen Staphylococcus aureus expresses a surface-exposed receptor, Iron surface determinant B (IsdB), that captures free human hemoglobin (Hb) and removes heme to retrieve iron, an essential nutrient for bacterial proliferation inside the host. Using single-particle cryo-electron microscopy, we solved the structure of two complexes between Hb and IsdB that represent snapshots of the initial interaction, where heme is still bound to Hb, and the final complex after completion of heme extraction. The structural and dynamic details unlocked through these structures will boost the design of inhibitors of IsdB:Hb interaction that might work as innovative antimicrobials.

The emergence of bacteria that co-express serine- and metallo- carbapenemases is a threat to the efficacy of the available β-lactam antibiotic armamentarium. The 4-amino-1,2,4-triazole-3-thione scaffold has been selected as the starting chemical moiety in the design of a small library of β-Lactamase inhibitors (BLIs) with extended activity profiles. The synthesised compounds have been validated in vitro against class A serine β−Lactamase (SBLs) KPC-2 and class B1 metallo β−Lactamases (MBLs) VIM-1 and IMP-1. Of the synthesised derivatives, four compounds showed cross-class micromolar inhibition potency and therefore underwent in silico analyses to elucidate their binding mode within the catalytic pockets of serine- and metallo-BLs. Moreover, several members of the synthesised library have been evaluated, in combination with meropenem (MEM), against clinical strains that overexpress BLs for their ability to synergise carbapenems.

In the search for new chemical scaffolds able to afford NLRP3 inflammasome inhibitors, we used a pharmacophore-hybridization strategy by combining the structure of the acrylic acid derivative INF39 with the 1-(piperidin-4-yl)1,3-dihydro-2H-benzo[d]imidazole-2-one substructure present in HS203873, a recently identified NLRP3 binder. A series of differently modulated benzo[d]imidazole-2-one derivatives were designed and synthesised. The obtained compounds were screened in vitro to test their ability to inhibit NLRP3-dependent pyroptosis and IL-1β release in PMA-differentiated THP-1 cells stimulated with LPS/ATP. The selected compounds were evaluated for their ability to reduce the ATPase activity of human recombinant NLRP3 using a newly developed assay. From this screening, compounds 9, 13 and 18, able to concentration-dependently inhibit IL-1β release in LPS/ATP-stimulated human macrophages, emerged as the most promising NLRP3 inhibitors of the series. Computational simulations were applied for building the first complete model of the NLRP3 inactive state and for identifying possible binding sites available to the tested compounds. The analyses led us to suggest a mechanism of protein–ligand binding that might explain the activity of the compounds.

α-amylases are ubiquitous enzymes belonging to the glycosyl hydrolase (GH13) family, whose members share a high degree of sequence identity, even between distant organisms. To understand the determinants of catalytic activity of α-amylases throughout evolution, and to investigate the use of homologous enzymes as a model for the human one, we compared human salivary α-amylase, Aspergillus oryzae α-amylase and pancreatic porcine α-amylase, using a combination of in vitro and in silico approaches. Enzyme sequences were aligned, and structures superposed, whereas kinetics were spectroscopically studied by using commercial synthetic substrates. These three enzymes show strikingly different activities, specifically mediated by different ions, despite relevant structural homology. Our study confirms that the function of α-amylases throughout evolution has considerably diverged, although key structural determinants, such as the catalytic triad and the calcium-binding pocket, have been retained. These functional differences need to be carefully considered when α-amylases, from different organisms, are used as a model for the human enzymes. In this frame, particular focus is needed for the setup of proper experimental conditions.

β-lactamases (BLs) represent the most frequent cause of antimicrobial resistance in Gram-negative bacteria. Despite the continuous efforts in the development of BL inhibitors (BLIs), new BLs able to hydrolyze the last developed antibiotics rapidly emerge. Moreover, the insurgence rate of effective mutations is far higher than the release of BLIs able to counteract them. This results in a shortage of antibiotics that is menacing the effective treating of infectious diseases. The situation is made even worse by the co-expression in bacteria of BLs with different mechanisms and hydrolysis spectra, and by the lack of inhibitors able to hit them all. Differently from other targets, BL flexibility has not been deeply exploited for drug design, possibly because of the small protein size, for their apparent rigidity and their high fold conservation. In this mini-review, we discuss the evidence for BL binding site dynamics being crucial for catalytic efficiency, mutation effect, and for the design of new inhibitors. Then, we report on identified allosteric sites in BLs and on possible allosteric inhibitors, as a strategy to overcome the frequent occurrence of mutations in BLs and the difficulty of competing efficaciously with substrates. Nevertheless, allosteric inhibitors could work synergistically with traditional inhibitors, increasing the chances of restoring bacterial susceptibility towards available antibiotics.