Bacterial structural genomics target enabled by a recently discovered potent fungal acetyl-CoA synthetase inhibitor.

The compound ethyl-adenosyl monophosphate ester (ethyl-AMP) has been shown to effectively inhibit acetyl-CoA synthetase (ACS) enzymes and to facilitate the crystallization of fungal ACS enzymes in various contexts. In this study, the addition of ethyl-AMP to a bacterial ACS from Legionella pneumophila resulted in the determination of a co-crystal structure of this previously elusive structural genomics target. The dual functionality of ethyl-AMP in both inhibiting ACS enzymes and promoting crystallization establishes its significance as a valuable resource for advancing structural investigations of this class of proteins.

Novel Keto-Alkyl-Pyridinium Antifungal Molecules Active in Models of In Vivo Candida albicans Vascular Catheter Infection and Ex Vivo Candida auris Skin Colonization.

New antifungal therapies are needed for both systemic, invasive infections in addition to superficial infections of mucosal and skin surfaces as well as biofilms associated with medical devices. The resistance of biofilm and biofilm-like growth phases of fungi contributes to the poor efficacy of systemic therapies to nonsystemic infections. Here, we describe the identification and characterization of a novel keto-alkyl-pyridinium scaffold with broad spectrum activity (2 to 16 µg/mL) against medically important yeasts and molds, including clinical isolates resistant to azoles and/or echinocandins. Furthermore, these keto-alkyl-pyridinium agents retain substantial activity against biofilm phase yeast and have direct activity against hyphal A. fumigatus. Although their toxicity precludes use in systemic infections, we found that the keto-alkyl-pyridinium molecules reduce Candida albicans fungal burden in a rat model of vascular catheter infection and reduce Candida auris colonization in a porcine ex vivo model. These initial preclinical data suggest that molecules of this class may warrant further study and development for nonsystemic applications.

Novel keto-alkyl-pyridinium antifungal molecules active in models of in vivo Candida albicans vascular catheter infection and ex vivo Candida auris skin colonization.

New antifungal therapies are needed for both systemic, invasive infections as well as superficial infections of mucosal and skin surfaces as well as biofilms associated with medical devices. The resistance of biofilm and biofilm-like growth phases of fungi contributes to the poor efficacy of systemic therapies to non-systemic infections. Here, we describe the identification and characterization of a novel keto-alkyl-pyridinium scaffold with broad spectrum activity (2-16 µg/mL) against medically important yeasts and moulds, including clinical isolates resistant to azoles and/or echinocandins. Furthermore, these keto-alkyl-pyridinium agents retain substantial activity against biofilm phase yeast and have direct activity against hyphal A. fumigatus . Although their toxicity precludes use in systemic infections, we found that the keto-alkyl-pyridinium molecules reduce C. albicans fungal burden in a rat model of vascular catheter infection and reduce Candida auris colonization in a porcine ex vivo model. These initial pre-clinical data suggest that molecules of this class may warrant further study and development.

Traditional Kenyan herbal medicine: exploring natural products' therapeutics against schistosomiasis.

Praziquantel (PZQ) remains the only drug of choice for the treatment of schistosomiasis, caused by parasitic flatworms. The widespread use of PZQ in schistosomiasis endemic areas for about four decades raises concerns about the emergence of resistance of Schistosoma spp. to PZQ under drug selection pressure. This reinforces the urgency in finding alternative therapeutic options that could replace or complement PZQ. We explored the potential of medicinal plants commonly used by indigenes in Kenya for the treatment of various ailments including malaria, pneumonia, and diarrhoea for their antischistosomal properties. Employing the Soxhlet extraction method with different solvents, seven medicinal plants Artemisia annua, Ajuga remota, Bredilia micranta, Cordia africana, Physalis peruviana, Prunus africana and Senna didymobotrya were extracted. Qualitative phytochemical screening was performed to determine the presence of various phytochemicals in the plant extracts. Extracts were tested against Schistosoma mansoni newly transformed schistosomula (NTS) and adult worms and the schistosomicidal activity was determined by using the adenosine triphosphate quantitation assay. Phytochemical analysis of the extracts showed different classes of compounds such as alkaloids, tannins, terpenes, etc., in plant extracts active against S. mansoni worms. Seven extracts out of 22 resulted in <20% viability against NTS in 24 h at 100 µg/ml. Five of the extracts with inhibitory activity against NTS showed >69.7% and ≥72.4% reduction in viability against adult worms after exposure for 24 and 48 h, respectively. This study provides encouraging preliminary evidence that extracts of Kenyan medicinal plants deserve further study as potential alternative therapeutics that may form the basis for the development of the new treatments for schistosomiasis.

Structural Characterization of the Reaction and Substrate Specificity Mechanisms of Pathogenic Fungal Acetyl-CoA Synthetases.

Acetyl CoA synthetases (ACSs) are Acyl-CoA/NRPS/Luciferase (ANL) superfamily enzymes that couple acetate with CoA to generate acetyl CoA, a key component of central carbon metabolism in eukaryotes and prokaryotes. Normal mammalian cells are not dependent on ACSs, while tumor cells, fungi, and parasites rely on acetate as a precursor for acetyl CoA. Consequently, ACSs have emerged as a potential drug target. As part of a program to develop antifungal ACS inhibitors, we characterized fungal ACSs from five diverse human fungal pathogens using biochemical and structural studies. ACSs catalyze a two-step reaction involving adenylation of acetate followed by thioesterification with CoA. Our structural studies captured each step of these two half-reactions including the acetyl-adenylate intermediate of the first half-reaction in both the adenylation conformation and the thioesterification conformation and thus provide a detailed picture of the reaction mechanism. We also used a systematic series of increasingly larger alkyl adenosine esters as chemical probes to characterize the structural basis of the exquisite ACS specificity for acetate over larger carboxylic acid substrates. Consistent with previous biochemical and genetic data for other enzymes, structures of fungal ACSs with these probes bound show that a key tryptophan residue limits the size of the alkyl binding site and forces larger alkyl chains to adopt high energy conformers, disfavoring their efficient binding. Together, our analysis provides highly detailed structural models for both the reaction mechanism and substrate specificity that should be useful in designing selective inhibitors of eukaryotic ACSs as potential anticancer, antifungal, and antiparasitic drugs.