Antonio De León Rodríguez

Fermentation of lactose and its constituent sugars by Escherichia coli WDHL: Impact on hydrogen production

Fermentations of lactose, glucose and galactose using Escherichia coli WDHL, a hydrogen over producer strain, were performed. With glucose as substrate pyruvate was mainly routed to the lactate pathway, resulting in hydrogen production and yield of 1037 mL and 0.30 mol H(2)/mol of glucose, respectively. When galactose was the substrate, the pyruvate formate lyase pathway was the main route for pyruvate and a fermentation yield of 1.12 mol H(2)/mol of galactose and a hydrogen production of 2080 mL were obtained. The fermentation of lactose or glucose plus galactose showed a similar yield of 1.02 mol H(2)/mol of hexose consumed. This work clearly demonstrated that the kinetics of hydrogen and metabolites production as well as the hydrogen yield were affected by the type of sugar used as substrate as reflected by the deviations from the metabolic hydrogen-production pathway.

Expansion of human hematopoietic stem cells for transplantation: trends and perspectives

Umbilical cord blood transplantation is clinically limited by its low progenitor cell content. Ex vivo expansion has become an alternative to increase the cell dose available for transplants. Expansion has been evaluated in several ways such as static cultures combining growth factors or mimicking the natural microenvironment using co-culture systems. However, static cultures have a small volume capacity and therefore large-scale expansion has been addressed using bioreactors. These and other biotechnological approaches for the expansion of hematopoietic progenitors and their utility to study several aspects of hematopoietic stem cell biology are discussed here.

Periplasmic expression and recovery of human interferon gamma in Escherichia coli

A synthetic human interferon gamma (hIFN-gamma) gene was fused to SP1 and SP3, two Sec-dependent artificial signal peptides to transport the hIFN-gamma to the periplasm of Escherichia coli BL21-SI. The processing efficiency of both SP1-hIFN-gamma and SP3-hIFN-gamma was dependent on the culture medium as well as the post-induction temperature. Both precursors were processed completely when cells were cultured using minimal medium and a post-induction temperature of 32.5 degrees C, and only the processed hIFN-gamma was detected. The SP3 signal peptide was more efficient than SP1 for the secretion of hIFN-gamma. Sixty percent of the total hIFN-gamma was secreted to the periplasm using the SP3 signal peptide and a post-induction temperature of 20 degrees C. Using Tris-sucrose-dithiothreitol (TSD) hypertonic buffer, the periplasmic soluble hINF-gamma was recovered with a purity of 85%.

Optimization of human interferon gamma production in Escherichia coli by response surface methodology

The production of human interferon gamma (hIFN-γ) using a synthetic gene in Escherichia coli BL21-SI was optimized by response surface methodology (RSM) and a Box-Behnken design. The process variables studied were temperature, bio-mass concentration at induction time and the NaCl concentration as inducer. According to the Box-Behnken design, a second order response function was developed. The optimal expression conditions were a temperature of 32.6°C, induction biomass of 0.31 g/L and 0.3 M NaCl in minimal medium. The model prediction for the maximum hIFN-γ production was 77.3 mg/L, which corresponded satisfactorily with the experimental data. The hIFN-γ concentration attained under optimized conditions was 13-times higher than that obtained using the non-optimized conditions. We conclude that RSM is an effective method for the optimization of recombinant protein expression using synthetic genes in E. coli.

Identification of differential expressed transcripts in cervical cancer of Mexican patients

The aim of this study was to identify the gene expression profile in biopsies of patients with cervical intraepithelial neoplasia (CIN) 1, CIN 2, CIN 3, and microinvasive cancer by suppression subtractive hybridization and Southern blotting. After analyzing 1,800 cDNA clones, we found 198 upregulated genes, 166 downregulated, and no significant change of gene expression in 86 clones (p = 0.005). These results were validated by Northern blot analysis (p = 0.0001) in the identification of 28 overexpressed and 7 downregulated transcripts. We observed a set of genes related to the Notch signaling pathway that may be involved in the transformation of cervical cells and in the development to malignancy. The differentially expressed genes may provide useful information about the molecular mechanisms involved in human cervical carcinoma and as diagnostic markers.