Dolores Hidalgo Cuna de la Independencia Nacional, Mexico
Dolores Hidalgo Cuna de la Independencia Nacional, Mexico

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PubMed | Colegio de Mexico, Hospital Infantil Of Mexico Federico Gomez, National Autonomous University of Mexico, University of Colima and 3 more.
Type: Journal Article | Journal: International journal of molecular sciences | Year: 2016

Glucose-6-phosphate dehydrogenase (G6PD) deficiency in humans causes severe disease, varying from mostly asymptomatic individuals to patients showing neonatal jaundice, acute hemolysis episodes or chronic nonspherocytic hemolytic anemia. In order to understand the effect of the mutations in G6PD gene function and its relation with G6PD deficiency severity, we report the construction, cloning and expression as well as the detailed kinetic and stability characterization of three purified clinical variants of G6PD that present in the Mexican population: G6PD Zacatecas (Class I), Vanua-Lava (Class II) and Viangchan (Class II). For all the G6PD mutants, we obtained low purification yield and altered kinetic parameters compared with Wild Type (WT). Our results show that the mutations, regardless of the distance from the active site where they are located, affect the catalytic properties and structural parameters and that these changes could be associated with the clinical presentation of the deficiency. Specifically, the structural characterization of the G6PD Zacatecas mutant suggests that the R257L mutation have a strong effect on the global stability of G6PD favoring an unstable active site. Using computational analysis, we offer a molecular explanation of the effects of these mutations on the active site.


PubMed | National Autonomous University of Mexico, Centro Mexicano Of Fomento Al Desarrollo, Laboratorio Of Bioquimica Genetica, Laboratorio Of Biologia Molecular and 2 more.
Type: Journal Article | Journal: Nutrients | Year: 2015

The importance of prebiotics consumption is increasing all over the world due to their beneficial effects on health. Production of better prebiotics from endemic plants raises possibilities to enhance nutritional effects in vulnerable population groups. Fructans derived from Agave Plant have demonstrated their safety and efficacy as prebiotics in animal models. Recently, the safety in humans of two fructans obtained from Agave tequilana (Metlin() and Metlos()) was demonstrated.This study aimed to demonstrate the efficacy as prebiotics of Metlin() and Metlos() in newborns of a randomized, double blind, controlled trial with a pilot study design. Biological samples were taken at 20 7 days, and three months of age from healthy babies. Outcomes of efficacy include impact on immune response, serum ferritin, C-reactive protein, bone metabolism, and gut bacteria changes.There were differences statistically significant for the groups of infants fed only with infant formula and with formula enriched with Metlin() and Metlos().Our results support the efficacy of Metlin() and Metlos() as prebiotics in humans, and stand the bases to recommend their consumption.ClinicalTrials.gov, NCT 01251783.


Chavez C.J.,Institute Investigaciones Biologicas | Ortega P.,Institute Investigaciones Biologicas | Leal J.,Institute Investigaciones Biologicas | D'Escrivan A.,University of Zulia | And 2 more authors.
Anales de Pediatria | Year: 2010

Introduction: Vitamin A deficiency (VAD) is a worldwide public health problem. Epidemiological studies of VAD prevalence have been conducted in individuals with chromosome load and genetic potential compared with the general population; however, there are few studies in patients with Down's syndrome (DS). The objective of this study was to determine the prevalence of VAD and analyse nutritional status in patients with DS. Methods: A prospective and cross-sectional study was performed, with 50 karyotypically normal (KN) individuals (10.4±3.7 years old) and 38 randomly selected patients with DS (8.2±4.1 years old). Serum retinol was determined by HPLC using the Bieri method, with an international reference standard to define VAD (serum retinol < 20 μg/dL). The data were analysed using the SAS/STAT statistical program. Results: The prevalence of VAD was 18.4% in individuals with DS and 4% in KN individuals (OR: 5.42; 95% CI=0.93 - 40.64; p=0.02). Children with DS between two and six years old shown a significativily lower serum retinol (p=< 0.05).The patients with DS also showed a significant decrease in height and weight compared to KN (p=< 0.001). Conclusions: The prevalence of VAD detected in patients with DS could be considered a public health problem. Also, the chromosome 21 trisomy represent a risk factor associated with VAD. © 2009 Asociación Española de Pediatría.


PubMed | Consejo Nacional de Ciencia y Tecnologia CONACYT, Laboratorio Of Nutricion Experimental, Autonomous University of Mexico City, Hospital Infantil Of Mexico Federico Gomez and 4 more.
Type: Review | Journal: International journal of molecular sciences | Year: 2016

Glucose-6-phosphate dehydrogenase (G6PD) is a key regulatory enzyme in the pentose phosphate pathway which produces nicotinamide adenine dinucleotide phosphate (NADPH) to maintain an adequate reducing environment in the cells and is especially important in red blood cells (RBC). Given its central role in the regulation of redox state, it is understandable that mutations in the gene encoding G6PD can cause deficiency of the protein activity leading to clinical manifestations such as neonatal jaundice and acute hemolytic anemia. Recently, an extensive review has been published about variants in the


PubMed | Laboratorio Of Bioquimica Genetica, Conacyt Research Fellow Instituto Nacional Of Pediatria, National Autonomous University of Mexico, Laboratorio Of Neurociencias and Colegio de Mexico
Type: Journal Article | Journal: International journal of molecular sciences | Year: 2015

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.


Reyes-Vivas H.,Laboratorio Of Bioquimica Genetica | De La Mora-De La Mora I.,Laboratorio Of Bioquimica Genetica | Castillo-Villanueva A.,Laboratorio Of Bioquimica Genetica | Yepez-Mulia L.,Centro Medico Nacional Siglo XXI | And 12 more authors.
Antimicrobial Agents and Chemotherapy | Year: 2014

Giardiasis is highly prevalent in the developing world, and treatment failures with the standard drugs are common. This work deals with the proposal of omeprazole as a novel antigiardial drug, focusing on a giardial glycolytic enzyme used to follow the cytotoxic effect at the molecular level. We used recombinant technology and enzyme inactivation to demonstrate the capacity of omeprazole to inactivate giardial triosephosphate isomerase, with no adverse effects on its human counterpart. To establish the specific target in the enzyme, we used single mutants of every cysteine residue in triosephosphate isomerase. The effect on cellular triosephosphate isomerase was evaluated by following the remnant enzyme activity on trophozoites treated with omeprazole. The interaction of omeprazole with giardial proteins was analyzed by fluorescence spectroscopy. The susceptibility to omeprazole of drug-susceptible and drug-resistant strains of Giardia lamblia was evaluated to demonstrate its potential as a novel antigiardial drug. Our results demonstrate that omeprazole inhibits giardial triosephosphate isomerase in a species-specific manner through interaction with cysteine at position 222. Omeprazole enters the cytoplasmic compartment of the trophozoites and inhibits cellular triosephosphate isomerase activity in a dose-dependent manner. Such inhibition takes place concomitantly with the cytotoxic effect caused by omeprazole on trophozoites. G. lamblia triosephosphate isomerase (GlTIM) is a cytoplasmic protein which can help analyses of how omeprazole works against the proteins of this parasite and in the effort to understand its mechanism of cytotoxicity. Our results demonstrate the mechanism of giardial triosephosphate isomerase inhibition by omeprazole and show that this drug is effective in vitro against drug-resistant and drug-susceptible strains of G. lamblia. Copyright © 2014, American Society for Microbiology. All Rights Reserved.


Gomez-Manzo S.,Laboratorio Of Bioquimica Genetica | Terron-Hernandez J.,Laboratorio Of Bioquimica Genetica | De la Mora-De la Mora I.,Laboratorio Of Bioquimica Genetica | Gonzalez-Valdez A.,National Autonomous University of Mexico | And 8 more authors.
International journal of molecular sciences | Year: 2014

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 k(cat) (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.


PubMed | Laboratorio Of Bioquimica Genetica, National Autonomous University of Mexico and CONACyT
Type: Journal Article | Journal: International journal of molecular sciences | Year: 2014

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 k(cat) (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.


PubMed | Laboratorio Of Bioquimica Genetica
Type: Journal Article | Journal: Biochimica et biophysica acta | Year: 2013

The deficiency of human triosephosphate isomerase (HsTIM) generates neurological alterations, cardiomyopathy and premature death. The mutation E104D is the most frequent cause of the disease. Although the wild type and mutant exhibit similar kinetic parameters, it has been shown that the E104D substitution induces perturbation of an interfacial water network that, in turn, reduces the association constant between subunits promoting enzyme inactivation. To gain further insight into the effects of the mutation on the structure, stability and function of the enzyme, we measured the sensitivity of recombinant E104D mutant and wild type HsTIM to limited proteolysis. The mutation increases the susceptibility to proteolysis as consequence of the loss of rigidity of its overall 3-D structure. Unexpectedly, it was observed that proteolysis of wild type HsTIM generated two different stable nicked dimers. One was formed in relatively short times of incubation with proteinase K; as shown by spectrometric and crystallographic data, it corresponded to a dimer containing a nicked monomer and an intact monomer. The formation of the other nicked species requires relatively long incubation times with proteinase K and corresponds to a dimer with two clipped subunits. The first species retains 50% of the original activity, whereas the second species is inactive. Collectively, we found that the E104D mutant is highly susceptible to proteolysis, which in all likelihood contributes to the pathogenesis of enzymopathy. In addition, the proteolysis data on wild type HsTIM illustrate an asymmetric conduct of the two monomers.

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