RESUMEN
Glioblastoma is one of the most lethal tumors, displaying striking cellular heterogeneity and drug resistance. The prognosis of patients suffering from glioblastoma after 5 years is only 5%. In the present work, capsaicin analogues bearing modifications on the acyl chain with long-chain fatty acids showed promising anti-tumoral activity by its cytotoxicity on U-87 and U-138 glioblastoma multiforme cells. The capsaicin analogues were enzymatically synthetized with cross-linked enzyme aggregates of lipase B from Candida antarctica (CALB). The catalytic performance of recombinant CALB-CLEAs was compared to their immobilized form on a hydrophobic support. After 72 h of reaction, the synthesis of capsaicin analogues from linoleic acid, docosahexaenoic acid, and punicic acid achieved a maximum conversion of 69.7, 8.3 and 30.3% with CALB-CLEAs, respectively. Similar values were obtained with commercial CALB, with conversion yields of 58.3, 24.2 and 22% for capsaicin analogues from linoleic acid, DHA and punicic acid, respectively. Olvanil and dohevanil had a significant cytotoxic effect on both U-87 and U-138 glioblastoma cells. Irrespective of the immobilization form, CALB is an efficient biocatalyst for the synthesis of anti-tumoral capsaicin derivatives. KEY POINTS: ⢠This is the first report concerning the enzymatic synthesis of capsaicin analogues from docosahexaenoic acid and punicic acid with CALB-CLEAs. ⢠The viability U-87 and U-138 glioblastoma cells was significantly affected after incubation with olvanil and dohevanil. ⢠Capsaicin analogues from fatty acids obtained by CALB-CLEAs are promising candidates for therapeutic use as cytotoxic agents in glioblastoma cancer cells.
Asunto(s)
Capsaicina , Glioblastoma , Humanos , Capsaicina/farmacología , Enzimas Inmovilizadas/metabolismo , Glioblastoma/tratamiento farmacológico , Proteínas Fúngicas/metabolismoRESUMEN
Biocatalytic processes play a crucial role in the valorization of lignin; therefore, methods enabling the monitoring of enzymes such as ß-etherases, capable of breaking ß-O-4 aryl-ether bonds, are of significant biotechnological interest. A novel method for quantifying ß-etherase activity was developed based on the ß-ester bond formation between a chromophore and acetovainillone. The chromogenic substrate ß-(ρ-nitrophenoxy)-α-acetovanillone (PNPAV), was chemically synthesized. Kintetic monitoring of ρ-nitrophenolate release at 410 nm over 10 min, using recombinant LigF from Sphingobium sp SYK-6, LigF-AB and LigE-AB from Althererytrobacter sp B11, yielded enzimatic activities of 404. 3 mU/mg, 72 mU/mg, and 50 mU/mg, respectively. This method is applicable in a pH range of 7.0-9.0, with a sensitivity of up to 50 ng of enzyme, exhibiting no interference with lipolytic, glycolytic, proteolytic, and oxidoreductase enzymes.
Asunto(s)
Compuestos Cromogénicos , Sphingomonadaceae , Oxidorreductasas/química , Proteínas Bacterianas/química , Lignina/químicaRESUMEN
Triacylglycerols (TAGs) are a primary energy source for marine mammals during lipid digestion. Walruses (Odobenus rosmarus divergens) consume prey with a high content of long-chain polyunsaturated fatty acids; however, their digestive physiology and lipid digestion remain poorly studied. The present study aims to model and characterize the gastric (PWGL) and pancreatic (PWPL) lipases of Pacific walruses using an in-silico approach. The confident 3D models of PWGL and PWPL were obtained via homology modeling and protein threading and displayed the structural features of lipases. Molecular docking analysis demonstrated substrate selectivity for long-chain TAG (Trieicosapentaenoin; TC20:5n-3) in PWGL and short-chain TAG (Trioctanoin; TC8:0) in PWPL. Molecular dynamics simulations demonstrate that PWGL bound to tridocosahexaenoin (TC22:6n-3), the protein is considerably stable at all three salinity conditions, but fluctuations are observed in the regions associated with catalytic sites and the lid, indicating the potential hydrolysis of the substrate. This is the first study to report on the digestion of TAGs in walruses, including modeling and lipases characterization and proposing a digestive tract for pinnipeds.
Asunto(s)
Lipasa , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Páncreas , Animales , Lipasa/metabolismo , Lipasa/química , Páncreas/enzimología , Morsas/metabolismo , Metabolismo de los Lípidos , Especificidad por Sustrato , Triglicéridos/metabolismo , Digestión , Estómago/enzimología , Secuencia de AminoácidosRESUMEN
The red palm weevil Rhynchophorus ferrugineus (Coleoptera: Curculionidae) is one of the most harmful pests for palm trees, causing serious economic damage worldwide. The present work aims to model and study the 3D structures of highly expressed odorant binding proteins from R. ferrugineus (RferOBPs) and identify possible binding modes and ligand release mechanism by docking and molecular dynamics. Highly confident 3D structures of a total of 11 odorant binding proteins (OBPs) were obtained with AlphaFold2. All 3D RferOBPs modeled structures displayed six characteristic α-helices, except for RfeOBP7 and RfeOBP10, which had an extra terminal α-helix. Among the eleven modeled RferOBPs, RferOBP4 was highly expressed in the antennae and subsequently selected for further analyses. Molecular docking analyses demonstrated that ferruginol, α-pinene, DEET, and picaridin can favorably bind the RferOBP4 cavity with low affinity energies. Molecular dynamic simulations of RferOBP4 bound to ferruginol at different pH values showed that low pH environments dictate a structural change into an apo-state that modifies the number of tunnels where the ligand can coexist, further triggering ligand release by a pH-dependent mechanism. This is the first report concerning the modelling and study of ligand binding modes and release mechanism of R. ferrugineus OBPs.Communicated by Ramaswamy H. Sarma.
RESUMEN
Halophilic archaea are capable of producing fructans, which are fructose-based polysaccharides. However, their biochemical characterization and biological and technological properties have been scarcely studied. The aim of this study was to evaluate the production, chemical characterization, biological and technological properties of a fructan inulin-type biosynthesized by a halophilic archaeon. Fructan extraction was performed through ethanol precipitation and purification by diafiltration. The chemical structure was elucidated using Fourier Transform-Infrared Spectroscopy and Nuclear Magnetic Resonance (NMR). Haloarcula sp. M1 biosynthesizes inulin with an average molecular weight of 8.37 × 106 Da. The maximal production reached 3.9 g of inulin per liter of culture within seven days. The glass transition temperature of inulin was measured at 138.85 °C, and it exhibited an emulsifying index of 36.47 %, which is higher than that of inulin derived from chicory. Inulin from Haloarcula sp. M1 (InuH) demonstrates prebiotic capacity. This study represents the first report on the biological and technological properties of inulin derived from halophilic archaea.
Asunto(s)
Cichorium intybus , Haloarcula , Inulina , Fructanos , EtanolRESUMEN
Jellyfish are economically important organisms in diverse countries, carnivorous organisms that consume various prey (crustaceans, mollusks, bivalves, etc.) and dissolved carbohydrates in marine waters. This study was focused on detecting and quantifying the activity of digestive glycosidases from the cannonball jellyfish (Stomolophus sp. 2) to understand carbohydrate digestion and its temporal-spatial variation. Twenty-three jellyfish gastric pouches were collected in 2015 and 2016 in the Gulf of California in three localities (Las Guásimas, Hermosillo, and Caborca). Nine samples were in intra-localities from Las Guásimas. Chitinase (Ch), ß-glucosidase (ß-glu), and ß-N-acetylhexosaminidase (ß-NAHA) were detected in the gastric pouches. However, cellulase, exoglucanase, α-amylase, polygalacturonase, xylanase, and κ-carrageenase were undetected. Detected enzymes showed halotolerant glycolytic activity (i = 0-4 M NaCl), optimal pH, and temperature at 5.0 and 30-50 °C, respectively. At least five ß-glucosidase and two ß-N-acetylhexosaminidase were detected using zymograms; however, the number of proteins with chitinase activity is not precise. The annual variation of cannonball jellyfish digestive glycosidases from Las Guásimas between 2015-2016 does not show significant differences despite the difference in phytoplankton measured as chlorophyll α (1.9 and 3.4 mg/m3, respectively). In the inter-localities, the glycosidase activity was statistically different in all localities, except for ß-N-acetylhexosaminidase activity between Caborca and Hermosillo (3,009.08 ± 87.95 and 3,101.81 ± 281.11 mU/g of the gastric pouch, respectively), with chlorophyll α concentrations of 2.6, 3.4 mg/m3, respectively. For intra-localities, the glycosidase activity did not show significant differences, with a mean chlorophyll α of 1.3 ± 0.1 mg/m3. These results suggest that digestive glycosidases from Stomolophus sp. 2 can hydrolyze several carbohydrates that may belong to their prey or carbohydrates dissolved in marine waters, with salinity over ≥ 0.6 M NaCl and diverse temperature (4-80 °C) conditions. Also, chlorophyll α is related to glycosidase activity in both seasons and inter-localities, except for chitinase activity in an intra-locality (Las Guásimas).
Asunto(s)
Celulasas , Quitinasas , Escifozoos , Animales , Glicósido Hidrolasas , Cloruro de Sodio , Escifozoos/química , beta-N-Acetilhexosaminidasas , Carbohidratos , ClorofilaRESUMEN
BACKGROUND: Arylacetamide deacetylase (AADAC) is a lipolytic enzyme involved in xenobiotic metabolism. The characterization in terms of activity and substrate preference has been limited to a few mammalian species. The potential role and catalytic activities of AADAC from other organisms are still poorly understood. Therefore, in this work, the physicochemical properties, proteomic analysis, and protein-protein interactions from Gnathostomata organisms were investigated. RESULTS: The analysis were performed with 142 orthologue sequences with ~ 48-100% identity with human AADAC. The catalytic motif HGG[A/G] tetrapeptide block was conserved through all AADAC orthologues. Four variations were found in the consensus pentapeptide GXSXG sequence (GDSAG, GESAG, GDSSG, and GSSSG), and a novel motif YXLXP was found. The prediction of N-glycosylation sites projected 4, 1, 6, and 4 different patterns for amphibians, birds, mammals, and reptiles, respectively. The transmembrane regions of AADAC orthologues were not conserved among groups, and variations in the number and orientation of the active site and C-terminal carboxyl were observed among the sequences studied. The protein-protein interaction of AADAC orthologues were related to cancer, lipid, and xenobiotic metabolism genes. CONCLUSION: The findings from this computational analysis offer new insight into one of the main enzymes involved in xenobiotic metabolism from mammals, reptiles, amphibians, and birds and its potential use in medical and veterinarian biotechnological approaches.
RESUMEN
Halophilic bacterias from saline soil from former Lake Texcoco were isolated, identified based on 16 rRNA and tested to produce glucolytic, nucleolytic, proteolytic and lipolytic exoenzymes. The Bacillus, Virgibacillus, Kocuria, Salinicoccus, Gracilibacillus, Halobacillus, Tenuibacillus and Nesterekonia genera where identified. Lipase/eserases and proteases from Nesterenkonia sp. and Nesterenkonia aethiopica showed halotolerant characteristics and were selected to synthesize the oleochemical n-butyl oleate and antioxidant peptides from muscle protein of common carp (Cyprinus carpio), respectively. In organic media (2,2,4-Trimethylpentane), the lipase/esterases from Nesterenkonia sp. (0.6 U/mL) and N. aethiopica (1.2 U/mL) achieved a 62.7% and 53.2% of n-butyl oleate conversion, respectively. The protein hydrolysis from muscle of common carp (C. carpio) showed a degree of hydrolysis of 4.5 ± 0.2% and 2.8 ± 0.1% when proteases from Nesterenkonia sp. and N. aethiopica were used, respectively. Three peptidic fractions ranging molecular masses between 254 and 1002 Da [M + H] show antioxidant scavenging activity, and the principal fraction with a peptide of 547.3 Da [M + H] showed an inhibition of 37.7 ± 1.8% and 16.3 ± 0.6%, when 2,2-diphenyl-1-picrylhydrazyl and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) were used, respectively. These findings showed that the enzymatic battery of the halophilic bacteria from former lake Texcoco can be used in hydrolysis and synthesis of molecules with applications in different fields as food technology or bioenergy.
Asunto(s)
Antioxidantes/metabolismo , Bacterias/clasificación , Bacterias/metabolismo , Ácidos Oléicos/metabolismo , Tolerancia a la Sal , Animales , Antioxidantes/química , Bacterias/genética , Bacterias/aislamiento & purificación , Proteínas Bacterianas/metabolismo , Carpas/metabolismo , Esterasas/metabolismo , Hidrólisis , Lagos , Lipasa/metabolismo , Péptido Hidrolasas/metabolismo , Péptidos/química , Péptidos/metabolismo , Filogenia , ARN Ribosómico 16S/genética , Microbiología del SueloRESUMEN
Pigments are present in a broad variety of terrestrial and aquatic organisms. The cannonball jellyfish (Stomolophus sp. 2) is an important fishery resource in the northwest of Mexico and is processed to be traded and consumed as seafood. During the process, water with a soluble blue pigment and other compounds are discarded to the environment. In this work, we present some properties of the blue pigment from Stomolophus sp.2 (S2bp), to decide if it could be considered as a potential value-added waste and avoid the blue proteinaceous pigment wastewater. S2bp was purified to homogeneity and had a molecular mass of 28.0 kDa; this protein exhibited a Êmax at 650 nm, contained Zn2+ and Cu2+ metal ions, and was stable from 10 to 50 °C and in a pH range of 3.0 to 13.0 for 1 h. It had halotolerant characteristics maintaining the blue coloration in a broad range of ionic strength (0-4 M NaCl) and showed changes in Êmax with chaotropic salts. In addition, S2bp was stable in the presence of organic acids and EDTA and in zwitterionic, anionic, and nonionic detergents at critical micellar concentration. However, oxidant reagents like NaClO and H2O2 decrease the coloration. These results show that the jellyfish pigment is a stable protein which makes it an alternative pigment for the food industry.