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Saúl Gómez-Manzo Jessica Terrón-Hernández Ignacio De la Mora-De la Mora Abigail González-Valdez Jaime Marcial-Quino Itzhel García-Torres America Vanoye-Carlo Gabriel López-Velázquez Gloria Hernández-Alcántara Jesús Oria-Hernández Horacio Reyes-Vivas Sergio Enríquez-Flores 《International journal of molecular sciences》2014,15(11):21179-21201
Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common enzyme deficiency worldwide, causing a wide spectrum of conditions with severity classified from the mildest (Class IV) to the most severe (Class I). To correlate mutation sites in the G6PD with the resulting phenotypes, we studied four naturally occurring G6PD variants: Yucatan, Nashville, Valladolid and Mexico City. For this purpose, we developed a successful over-expression method that constitutes an easier and more precise method for obtaining and characterizing these enzymes. The kcat (catalytic constant) of all the studied variants was lower than in the wild-type. The structural rigidity might be the cause and the most evident consequence of the mutations is their impact on protein stability and folding, as can be observed from the protein yield, the T50 (temperature where 50% of its original activity is retained) values, and differences on hydrophobic regions. The mutations corresponding to more severe phenotypes are related to the structural NADP+ region. This was clearly observed for the Classes III and II variants, which became more thermostable with increasing NADP+, whereas the Class I variants remained thermolabile. The mutations produce repulsive electric charges that, in the case of the Yucatan variant, promote increased disorder of the C-terminus and consequently affect the binding of NADP+, leading to enzyme instability. 相似文献
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Saúl Gómez-Manzo José E. Escamilla Abigail González-Valdez Gabriel López-Velázquez América Vanoye-Carlo Jaime Marcial-Quino Ignacio de la Mora-de la Mora Itzhel Garcia-Torres Sergio Enríquez-Flores Martha Lucinda Contreras-Zentella Roberto Arreguín-Espinosa Peter M. H. Kroneck Martha Elena Sosa-Torres 《International journal of molecular sciences》2015,16(1):1293-1311
Gluconacetobacter diazotrophicus is a N2-fixing bacterium endophyte from sugar cane. The oxidation of ethanol to acetic acid of this organism takes place in the periplasmic space, and this reaction is catalyzed by two membrane-bound enzymes complexes: the alcohol dehydrogenase (ADH) and the aldehyde dehydrogenase (ALDH). We present strong evidence showing that the well-known membrane-bound Alcohol dehydrogenase (ADHa) of Ga. diazotrophicus is indeed a double function enzyme, which is able to use primary alcohols (C2–C6) and its respective aldehydes as alternate substrates. Moreover, the enzyme utilizes ethanol as a substrate in a reaction mechanism where this is subjected to a two-step oxidation process to produce acetic acid without releasing the acetaldehyde intermediary to the media. Moreover, we propose a mechanism that, under physiological conditions, might permit a massive conversion of ethanol to acetic acid, as usually occurs in the acetic acid bacteria, but without the transient accumulation of the highly toxic acetaldehyde. 相似文献
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Saúl Gómez-Manzo Jaime Marcial-Quino America Vanoye-Carlo Sergio Enríquez-Flores Ignacio De la Mora-De la Mora Abigail González-Valdez Itzhel García-Torres Víctor Martínez-Rosas Edgar Sierra-Palacios Fernando Lazcano-Pérez Eduardo Rodríguez-Bustamante Roberto Arreguin-Espinosa 《International journal of molecular sciences》2015,16(12):28657-28668
Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common enzymopathy in the world. More than 160 mutations causing the disease have been identified, but only 10% of these variants have been studied at biochemical and biophysical levels. In this study we report on the functional and structural characterization of three naturally occurring variants corresponding to different classes of disease severity: Class I G6PD Durham, Class II G6PD Santa Maria, and Class III G6PD A+. The results showed that the G6PD Durham (severe deficiency), and the G6PD Santa Maria and A+ (less severe deficiency) (Class I, II and III, respectively) affect the catalytic efficiency of these enzymes, are more sensitive to temperature denaturing, and affect the stability of the overall protein when compared to the wild type WT-G6PD. In the variants, the exposure of more and buried hydrophobic pockets was induced and monitored with 8-Anilinonaphthalene-1-sulfonic acid (ANS) fluorescence, directly affecting the compaction of structure at different levels and probably reducing the stability of the protein. The degree of functional and structural perturbation by each variant correlates with the clinical severity reported in different patients. 相似文献
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Jose Garcia-Torres 《Electrochimica acta》2010,55(20):5760-17971
A study of the best conditions to prepare smooth heterogeneous Co-Ag films with low amounts of S from a thiourea-based electrolytic bath has been performed. Using a 0.01 M AgClO4 + 0.1 M Co(ClO4)2 + 0.1 M thiourea + 0.1 M sodium gluconate + 0.3 M H3BO3 + 0.1 M NaClO4 bath, low temperature (10 °C) allowed obtaining compact and smooth deposits containing 2 wt.% sulphur. Decreasing thiourea content 0.06 M and increasing gluconate concentration up to 0.3 M, better deposits (more compact with lower sulphur content (1.2 wt.%)) were obtained. A clear influence of the species present in the bath on the film quality was observed: while gluconate favoured film cohesion, boric acid hindered hydrogen adsorption. For all films, fcc-Ag, hcp-Co and hcp-CoAg3 phases were always detected by XRD, TEM and electron diffraction, their proportions varying with the electrodeposition conditions. Magnetic measurements revealed that the increase in the CoAg3 led to an increase in the film coercivity. GMR values were only measured at cryogenic temperatures, they being higher for the deposits with the lowest sulphur content revealing that sulphur exerts a negative effect on magnetoresistance. 相似文献
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Cynthia Fernndez-Lainez Ignacio de la Mora-de la Mora Itzhel García-Torres Sergio Enríquez-Flores Luis A. Flores-Lpez Pedro Gutirrez-Castrelln Lilian Ypez-Mulia Felix Matadamas-Martínez Paul de Vos Gabriel Lpez-Velzquez 《International journal of molecular sciences》2021,22(17)
Giardiasis represents a latent problem in public health due to the exceptionally pathogenic strategies of the parasite Giardia lamblia for evading the human immune system. Strains resistant to first-line drugs are also a challenge. Therefore, new antigiardial therapies are urgently needed. Here, we tested giardial arginine deiminase (GlADI) as a target against giardiasis. GlADI belongs to an essential pathway in Giardia for the synthesis of ATP, which is absent in humans. In silico docking with six thiol-reactive compounds was performed; four of which are approved drugs for humans. Recombinant GlADI was used in enzyme inhibition assays, and computational in silico predictions and spectroscopic studies were applied to follow the enzyme’s structural disturbance and identify possible effective drugs. Inhibition by modification of cysteines was corroborated using Ellman’s method. The efficacy of these drugs on parasite viability was assayed on Giardia trophozoites, along with the inhibition of the endogenous GlADI. The most potent drug against GlADI was assayed on Giardia encystment. The tested drugs inhibited the recombinant GlADI by modifying its cysteines and, potentially, by altering its 3D structure. Only rabeprazole and omeprazole decreased trophozoite survival by inhibiting endogenous GlADI, while rabeprazole also decreased the Giardia encystment rate. These findings demonstrate the potential of GlADI as a target against giardiasis. 相似文献
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