RESUMO
Canavan disease is caused by mutations in the ASPA gene, leading to diminished catalytic activity of aspartoacylase in the brain. Clinical missense mutations are found throughout the enzyme structure, with many of these mutated enzymes having not only decreased activity but also compromised stability. High-throughput screening of a small molecule library has identified several compounds that significantly increase the thermal stability of the E285A mutant enzyme, the most predominant clinical mutation in Canavan disease, while having a negligible effect on the native enzyme. Based on the initial successes, some structural analogs of these initial hits were selected for further examination. Glutathione, NAAG and patulin were each confirmed to be competitive inhibitors, indicating the binding of these compounds at the dimer interface or near the active site of the E285A enzyme. The experimental results were theoretically examined with the help of the docking analysis method. The structure activity-guided optimization of these compounds can potentially lead to potential pharmacological chaperones that could alleviate the detrimental effect of ASPA mutations in Canavan patients.
RESUMO
Glucose, the primary substrate for ATP synthesis, is catabolized during glycolysis to generate ATP and precursors for the synthesis of other vital biomolecules. Opportunistic viruses and cancer cells often hijack this metabolic machinery to obtain energy and components needed for their replication and proliferation. One way to halt such energy-dependent processes is by interfering with the glycolytic pathway. 2-Deoxy-d-glucose (2-DG) is a synthetic glucose analogue that can inhibit key enzymes in the glycolytic pathway. The efficacy of 2-DG has been reported across an array of diseases and disorders, thereby demonstrating its broad therapeutic potential. Recent approval of 2-DG in India as a therapeutic approach for the management of the COVID-19 pandemic has brought renewed attention to this molecule. The purpose of this perspective is to present updated therapeutic avenues as well as a variety of chemical synthetic strategies for this medically useful sugar derivative, 2-DG.
Assuntos
Antivirais/uso terapêutico , Tratamento Farmacológico da COVID-19 , Desoxiglucose/química , Trifosfato de Adenosina/metabolismo , Antivirais/química , Antivirais/metabolismo , Antivirais/farmacologia , COVID-19/diagnóstico , COVID-19/virologia , Desoxiglucose/metabolismo , Desoxiglucose/farmacologia , Desoxiglucose/uso terapêutico , Epilepsia/diagnóstico , Epilepsia/tratamento farmacológico , Epilepsia/patologia , Glicólise/efeitos dos fármacos , Humanos , Marcação por Isótopo , Mitocôndrias/metabolismo , Neoplasias/diagnóstico , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Tomografia por Emissão de Pósitrons , SARS-CoV-2/isolamento & purificação , SARS-CoV-2/fisiologia , Relação Estrutura-Atividade , Replicação Viral/efeitos dos fármacosRESUMO
Today, the world is suffering from the pandemic of a novel coronavirus disease (COVID-19), a respiratory illness caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). This pandemic is the third fatal coronavirus outbreak that has already occurred in the 21st century. Even six months after its emergence, hundreds of thousands of people are still being infected with SARS-CoV-2, and thousands of lives are lost every day across the world. No effective therapy has been approved to date for the treatment of this disease, suggesting the need to broaden the scope in the search for effective treatments. Throughout history, folk medicine has been successfully used to treat various ailments in humans, and Traditional Chinese Medicine has been instrumental in the containment of a number of viral diseases. Owing to their high chemical diversity and safety profiles, natural products offer great promises as potentially effective antiviral drugs. In recent years, a large number of anti-coronaviral phytochemicals with different mechanisms of action have been identified. Among them, tetra-O-galloyl-ß-D-glucose, caffeic acid, and saikosaponin B2 block viral entry. A number of flavonoids inhibit viral proteases. Silvestrol inhibits protein synthesis. Myricetin and scutellarein inhibit viral replication. Emodin, luteolin, and quercetin demonstrate anti-coronaviral activity by inhibiting multiple processes in the virus life cycle. In this review, we critically evaluate the findings of the natural product-based anticoronaviral research that has been published during the last two decades, and attempt to provide a comprehensive description about their utility as potential broad-spectrum anti-coronaviral drugs, examining leads that may guide/facilitate anti-SARS-CoV-2 drug development studies.
Assuntos
Produtos Biológicos , COVID-19 , Antivirais/farmacologia , Antivirais/uso terapêutico , Produtos Biológicos/farmacologia , Humanos , Pandemias , SARS-CoV-2RESUMO
BACKGROUND: Diabetes mellitus type 2 (DMT2) is a leading metabolic disorder in the world. Anti-diabetic actions of phytochemicals from various medicinal herbs have been explored as an alternative therapy in the management of DMT2 due to adverse effects of synthetic drugs used in allopathic medicine. α-amylase and α-glucosidase inhibitory potential and phytochemical profiling were investigated in aqueous extracts of two new Cinnamomum zeylanicum accessions, namely C. zeylanicum Sri Wijaya (SW), C. zeylanicum Sri Gemunu (SG) and commercially available C. zeylanicum (CC). METHODS: Microwave Digestion (MD), Pressurized Water Extraction (PWE), Steam Distillation (SD), Solvent Extraction (SE), Decoction Water Extraction (DWE) and Infusion Water Extraction (IWE) methods were used to prepare Cinnamon quill extracts. Total phenolic content (TPC, Folin-Ciocalteu method) and Proanthocyanidin content (PC, vanillin assay), α-amylase and α-glucosidase inhibition of Cinnamon extracts were determined spectrophotometrically. The α-amylase and α-glucosidase inhibition were reported in terms of IC50 value. The phytochemical profiling was accomplished by GC-MS technique. RESULTS AND DISCUSSION: Lowest IC50 values were observed in PWE and DWE of SW. The highest PC and TPC were also observed in PWE and DWE of SW. Pressured water and decoctions are promising methods for the extraction of antidiabetic constituents from cinnamon. Benzoic acid, cinnamyl alcohol, benzyl alcohol, and 4-Allyl-2,6-dimethoxyphenol were identified as major compounds in SW extracts. These compounds are believed to be responsible for strong enzyme inhibitory activity of the extracts. CONCLUSIONS: This is the first study to explore the use of pressured and decoctions water to extract anti-diabetic phytochemicals from cinnamon. The extensive metabolite profiling of novel SW and SG extracts and comparison of that with commercially available CC are reported for the first time in this study. The C. zeylanicum, SW accession holds some promise in the management of diabetes.
RESUMO
S-Adenosyl-l-methionine (AdoMet) is an essential metabolite, serving in a very wide variety of metabolic reactions. The enzyme that produces AdoMet from l-methionine and ATP (methionine adenosyltransferase, MAT) is thus an attractive target for antimicrobial agents. We previously showed that a variety of methionine analogues are MAT substrates, yielding AdoMet analogues that function in specific methyltransfer reactions. However, this left open the question of whether the modified AdoMet molecules could support bacterial growth, meaning that they functioned in the full range of essential AdoMet-dependent reactions. The answer matters both for insight into the functional flexibility of key metabolic enzymes, and for drug design strategies for both MAT inhibitors and selectively toxic MAT substrates. In this study, methionine analogues were converted in vitro into AdoMet analogues, and tested with an Escherichia coli strain lacking MAT (ΔmetK) but that produces a heterologous AdoMet transporter. Growth that yields viable, morphologically normal cells provides exceptionally robust evidence that the analogue functions in every essential reaction in which AdoMet participates. Overall, the S-adenosylated derivatives of all tested l-methionine analogues modified at the carboxyl moiety, and some others as well, showed in vivo functionality sufficient to allow good growth in both rich and minimal media, with high viability and morphological normality. As the analogues were chosen based on incompatibility with the reactions via which AdoMet is used to produce acylhomoserine lactones (AHLs) for quorum sensing, these results support the possibility of using this route to selectively interfere with AHL biosynthesis without inhibiting bacterial growth.
Assuntos
Escherichia coli/crescimento & desenvolvimento , Escherichia coli/metabolismo , S-Adenosilmetionina/metabolismo , Acil-Butirolactonas/metabolismo , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Metionina Adenosiltransferase/genética , Metionina Adenosiltransferase/metabolismo , Estrutura Molecular , S-Adenosilmetionina/análogos & derivadosRESUMO
Canavan disease (CD) is a fatal, childhood neurological disorder caused by mutations in the ASPA gene, leading to catalytic deficiencies in the aspartoacylase (ASPA) enzyme and impaired N-acetyl-l-aspartic acid metabolism in the brain. To study the possible structural defects triggered by these mutations, four ASPA missense mutations associated with different disease severities have been structurally characterized. These mutant enzymes each have overall structures similar to that of the native ASPA enzyme, but with varying degrees of alterations that offer explanations for the respective loss of catalytic activity. The K213E mutant, a nonconservative mutant associated with a mild disease phenotype, has minimal structural differences compared to the native enzyme. In contrast, the loss of van der Waals contacts in the F295S mutant and the loss of hydrophobic and hydrogen bonding interactions in the Y231C mutant lead to a local collapse of the hydrophobic core structure in the carboxyl-terminal domain, contributing to a decrease in protein stability. The structure of the E285A mutant, the most common clinical mutant, reveals that the loss of hydrogen bonding interactions with the carboxylate side chain of Glu285 disturbs the active site architecture, leading to altered substrate binding and lower catalytic activity. Our improved understanding of the nature of these structural defects provides a basis for the development of treatment therapies for CD.
Assuntos
Amidoidrolases/química , Amidoidrolases/deficiência , Doença de Canavan/enzimologia , Domínio Catalítico/genética , Mutação de Sentido Incorreto/genética , Amidoidrolases/genética , Doença de Canavan/genética , Linhagem Celular , Cristalografia por Raios X , Humanos , Relação Estrutura-AtividadeRESUMO
Bacteria use quorum sensing to probe and respond to population densities in their external environment. The detection of quorum signaling molecules causes a virulence response in many pathogenic bacteria. Blocking this signaling pathway, without interfering with critical metabolic functions, would produce compounds that can disarm pathogens without killing them. By not blocking growth per se, this therapeutic approach would have a lower associated risk for the development of bacterial resistance. Modified forms of l-methionine can yield analogues of the essential methyl donor, S-adenosyl-l-methionine (AdoMet), by serving as substrates for AdoMet synthetase [Zano, S., et al. (2013) Arch. Biochem. Biophys. 536, 64]. The AdoMet analogues examined here were chosen for their putative inability to serve as precursors for the synthesis of the acylhomoserine lactone class of quorum sensing molecules. We now show that these AdoMet analogues can still function as methyl donors, for methylation of both DNA and catechol-based neurotransmitters. The rates of methyl transfer for several of these altered AdoMet analogues are comparable to those observed with unmodified AdoMet. Additional refinement of these structures is expected to produce lead compounds to be tested as selective therapeutic agents against infections by a broad range of pathogenic Gram-negative bacteria.
Assuntos
Metionina Adenosiltransferase/metabolismo , Metilação de DNA , Percepção de Quorum , S-Adenosilmetionina/metabolismoRESUMO
Canavan disease (CD) is a fatal neurological disorder caused by defects in the gene that encodes for a critical metabolic enzyme. The enzyme aspartoacylase catalyzes the deacetylation of N-acetylaspartate to produce acetate required for fatty acid biosynthesis in the brain. The loss of aspartoacylase activity leads to the demyelination and disrupted brain development that is found in CD patients. Sixteen different clinical mutants of aspartoacylase have been cloned, expressed and purified to examine their properties and the relationship between enzyme properties and disease phenotype. In contrast to numerous cell culture studies that reported virtually complete loss of function, each of these purified mutant enzymes was found to have measureable catalytic activity. However, the activities of these mutants are diminished, by as little as three-fold to greater than 100-fold when compared to the native enzyme. Many of these mutated enzyme forms show decreased thermal stability and an increased propensity for denaturation upon exposure to urea, but only four of the 16 mutants examined showed both diminished thermal and diminished conformational stability. Significantly, each of these lower stability mutants are responsible for the more severe phenotypes of CD, while patients with milder forms of CD have aspartoacylase mutants with generally high catalytic activity and with either good thermal or good conformational stability. These results suggest that the loss of catalytic function and the accumulation of N-acetylaspartate in Canavan disease is at least partially a consequence of the decreased protein stability caused by these mutations.