Pau Ferrer - Academia.edu (original) (raw)

Papers by Pau Ferrer

Pharmaceutics

Currently, three human papillomavirus (HPV) vaccines are already licensed and all of them are bas... more Currently, three human papillomavirus (HPV) vaccines are already licensed and all of them are based on virus-like particles (VLPs) of HPV L1 capsid protein. While about 38.0 million people were living with HIV in 2019, only 68% of HIV-infected individuals were accessing antiretroviral therapy as of the end of June 2020. Therefore, safe, effective, and affordable vaccines against those two viruses are immediately needed. Both HPV and HIV are sexually transmitted infections and one of the main access routes is the mucosal genital tract. Thus, the development of a combined vaccine that would protect against HPV and HIV infections is a logical effort in the fight against these two major global pathogens. In this study, a recombinant Pichia pastoris producing chimeric HPV-HIV L1P18 protein intracellularly was constructed. After cell disruption, the supernatant was collected, and the VLPs were purified by a combination of ammonium sulfate precipitation, size exclusion chromatography, ultr...

Methods in Molecular Biology

Yeasts are efficient cell factories and are commonly used for the production of recombinant prote... more Yeasts are efficient cell factories and are commonly used for the production of recombinant proteins for biopharmaceutical and industrial purposes. For such products high levels of correctly folded proteins are needed, which sometimes requires improvement and engineering of the expression system. The article summarizes major breakthroughs that led to the efficient use of yeasts as production platforms and reviews bottlenecks occurring during protein production. Special focus is given to the metabolic impact of protein production. Furthermore, strategies that were shown to enhance secretion of recombinant proteins in different yeast species are presented.

Miscanthussp. is regarded as suitable biomass for different biorefinery value chains. However, du... more Miscanthussp. is regarded as suitable biomass for different biorefinery value chains. However, due to high recalcitrance, its wide use is yet untapped. Termite is the most efficient lignocellulose degrading insect, and its success results from synergistic cooperation with its gut microbiome. Here, we investigated at holobiont level the dynamic adaptation of a higher termiteCortaritermessp. to imposedMiscanthusdiet, with a long-term objective of overcoming lignocellulose recalcitrance. We used an integrative omics approach, comprising amplicon sequencing, metagenomics and metatranscriptomics that we combined with enzymatic characterisation of carbohydrate active enzymes from termite gut Fibrobacteres and Spirochaetae. Adaptation to the new diet was evidenced by reduced gut bacterial diversity and modified gene expression profiles, further suggesting a shift towards utilisation of cellulose and arabinoxylan, two main components ofMiscanthuslignocellulose. Low identity of reconstructed...

Aids Reviews

Virus-like particles (VLPs) are a type of subunit vaccine which resembles viruses but do not cont... more Virus-like particles (VLPs) are a type of subunit vaccine which resembles viruses but do not contain any genetic material so that they are not infectious. VLPs maintain the same antigenic conformation to the original virus, and they could be a better vaccine candidate than live-attenuated and inactivated vaccines. In addition, compared to other subunit vaccines such as soluble protein, VLPs can stimulate both innate and adaptive immune responses effectively and safely against several pathogens by the closer morphology to its native virus. They have already been licensed as vaccines against Hepatitis B virus, human papillomavirus (HPV), and several veterinary diseases. Moreover, it has been investigated to prevent other viral infections including HIV. While HIV VLP-based vaccines have been studied over 35 years, none of them has been successful enough to reach even Phase III clinical trials. In this review, we summarize: (i) general features of VLPs; (ii) epidemiological data and current status of vaccine research and development on HPV and HIV; and (iii) previous studies held on HPV VLPs, HIV VLPs, and chimeric HPV/HIV VLPs including production methods and different animal immunization assays. Furthermore, we review present state of human clinical trials with VLPs and consider the potential to develop a successful preventive HIV vaccine using HPV VLP models. Finally, we discuss the benefits, limitations, and challenges of developing chimeric VLP-based HPV/HIV vaccines with recent findings, critical issues to improve VLP-based vaccines, and hot topics for the next 5 years to join the global effort to fight against these two pathogens.

Frontiers in Microbiology

Antimicrobial peptides (AMPs) have potent and durable antimicrobial activity to a wide range of f... more Antimicrobial peptides (AMPs) have potent and durable antimicrobial activity to a wide range of fungi and bacteria. The growing problem of drug-resistant pathogenic microorganisms, together with the lack of new effective compounds, has stimulated interest in developing AMPs as anti-infective molecules. PAF102 is an AMP that was rationally designed for improved antifungal properties. This cell penetrating peptide has potent and specific activity against major fungal pathogens. Cecropin A is a natural AMP with strong and fast lytic activity against bacterial and fungal pathogens, including multidrug resistant pathogens. Both peptides, PAF102 and Cecropin A, are alternative antibiotic compounds. However, their exploitation requires fast, cost-efficient production systems. Here, we developed an innovative system to produce AMPs in Pichia pastoris using the oleosin fusion technology. Oleosins are plant-specific proteins with a structural role in lipid droplet formation and stabilization, which are used as carriers for recombinant proteins to lipid droplets in plant-based production systems. This study reports the efficient production of PAF102 in P. pastoris when fused to the rice plant Oleosin 18, whereas no accumulation of Cecropin A was detected. The Ole18-PAF102 fusion protein targets the lipid droplets of the heterologous system where it accumulates to high levels. Interestingly, the production of this fusion protein induces the formation of lipid droplets in yeast cells, which can be additionally enhanced by the coexpression of a diacylglycerol transferase gene that allows a threefold increase in the production of the fusion protein. Using this high producer strain, PAF102 reaches commercially relevant yields of up to 180 mg/l of yeast culture. Moreover, the accumulation of PAF102 in the yeast lipid droplets facilitates its downstream extraction and recovery by flotation on density gradients, with the recovered PAF102 being biologically active against pathogenic fungi. Our results demonstrate that plant oleosin fusion technology can be transferred to the well-established P. pastoris cell factory to produce the PAF102 antifungal peptide, and potentially other AMPs, for multiple applications in crop protection, food preservation and animal and human therapies.

Applied and Environmental Microbiology

Increased hydrolysis of easily digestible biomass may lead to acidosis of anaerobic reactors and ... more Increased hydrolysis of easily digestible biomass may lead to acidosis of anaerobic reactors and decreased methane production. Previously, it was shown that the structure of microbial communities changed during acidosis, however once the conditions are back to optimal, biogas (initially CO2) production quickly restarts. This suggests the retention of the community functional redundancy during the process failure. In this study, with the use of metagenomics and downstream bioinformatics analyses, we characterize the carbohydrate hydrolytic potential of the microbial community, with a special focus on acidosis. To that purpose, carbohydrate active enzymes were identified and to further link the community hydrolytic potential with key microbes, bacterial genomes were re-constructed. In continuation, we characterized biochemically the specificity and activity of selected enzymes, thus verifying the accuracy of their in silico predictions. The results confirm the retention of the communi...

Biotechnology and Bioengineering

The methanol‐regulated alcohol oxidase promoter (PAOX1) of Pichia pastoris (syn. Komagataella spp... more The methanol‐regulated alcohol oxidase promoter (PAOX1) of Pichia pastoris (syn. Komagataella spp. ) is one of the strongest promoters for heterologous gene expression. Although increasing the gene dosage is a common strategy to improve recombinant protein productivities, P. pastoris strains harboring more than two copies of a Rhizopus oryzae lipase gene (ROL) have previously shown a decrease in cell growth, lipase production, and substrate consumption, as well as a significant transcriptional downregulation of methanol metabolism. This pointed to a potential titration effect of key transcriptional factors methanol expression regulator 1 (Mxr1) and methanol‐induced transcription factor (Mit1) regulating methanol metabolism caused by the insertion of multiple expression vectors. To prove this hypothesis, a set of strains carrying one and four copies of ROL (1C and 4C, respectively) were engineered to coexpress one or two copies of MXR1*, coding for an Mxr1 variant insensitive to repression by 14‐3‐3 regulatory proteins, or one copy of MIT1. Small‐scale cultures revealed that growth, Rol productivity, and methanol consumption were improved in the 4C‐MXR1* and 4C‐MIT1, strains growing on methanol as a sole carbon source, whereas only a slight increase in productivity was observed for re‐engineered 1C strains. We further verified the improved performance of these strains in glycerol‐/methanol‐limited chemostat cultures.

Microbial cell factories, Jan 22, 2018

Azotobacter vinelandii is a bacterium that produces alginate and polyhydroxybutyrate (P3HB); howe... more Azotobacter vinelandii is a bacterium that produces alginate and polyhydroxybutyrate (P3HB); however, the role of NAD(P)H/NAD(P)+ ratios on the metabolic fluxes through biosynthesis pathways of these biopolymers remains unknown. The aim of this study was to evaluate the NAD(P)H/NAD(P) + ratios and the metabolic fluxes involved in alginate and P3HB biosynthesis, under oxygen-limiting and non-limiting oxygen conditions. The results reveal that changes in the oxygen availability have an important effect on the metabolic fluxes and intracellular NADPH/NADP+ ratio, showing that at the lowest OTR (2.4 mmol L-1 h-1), the flux through the tricarboxylic acid (TCA) cycle decreased 27.6-fold, but the flux through the P3HB biosynthesis increased 6.6-fold in contrast to the cultures without oxygen limitation (OTR = 14.6 mmol L-1 h-1). This was consistent with the increase in the level of transcription of phbB and the P3HB biosynthesis. In addition, under conditions without oxygen limitation, the...

Quaderns D Italia, 2002

... to analogico scivoliamo nel triviale, d'ap-pendice. ... deseo.» Ha, senz'al... more ... to analogico scivoliamo nel triviale, d'ap-pendice. ... deseo.» Ha, senz'altro, ragione Luis Anto-nio de Villena: Penna è forse l'unico poeta del '900 italiano che ha voluto affidare la Sandro PENA Algo de fiebre Traducció di Luis Antonio de Villena, Valencia: Pre-textos, 2002

PLOS ONE, 2016

Motivation Genome-scale metabolic models (GEMs) are tools that allow predicting a phenotype from ... more Motivation Genome-scale metabolic models (GEMs) are tools that allow predicting a phenotype from a genotype under certain environmental conditions. GEMs have been developed in the last ten years for a broad range of organisms, and are used for multiple purposes such as discovering new properties of metabolic networks, predicting new targets for metabolic engineering, as well as optimizing the cultivation conditions for biochemicals or recombinant protein production. Pichia pastoris is one of the most widely used organisms for heterologous protein expression. There are different GEMs for this methylotrophic yeast of which the most relevant and complete in the published literature are iPP668, PpaMBEL1254 and iLC915. However, these three models differ regarding certain pathways, terminology for metabolites and reactions and annotations. Moreover, GEMs for some species are typically built based on the reconstructed models of related model organisms. In these cases, some organism-specific pathways could be missing or misrepresented. Results In order to provide an updated and more comprehensive GEM for P. pastoris, we have reconstructed and validated a consensus model integrating and merging all three existing models. In this step a comprehensive review and integration of the metabolic pathways included in each one of these three versions was performed. In addition, the resulting iMT1026 model includes a new description of some metabolic processes. Particularly new information described in recently published literature is included, mainly related to fatty acid and sphingolipid metabolism, glycosylation and cell energetics. Finally the reconstructed model was tested and validated, by comparing the results of the simulations with available empirical physiological datasets results obtained from a wide range of experimental PLOS ONE |

Farmacia Hospitalaria, 2005

Evaluar la calidad del proceso de llenado de carros en un sistema de distribución en dosis unitar... more Evaluar la calidad del proceso de llenado de carros en un sistema de distribución en dosis unitarias tras la implantación de un protocolo. Método: Se definieron cinco criterios de calidad: cuatro relativos a la medicación dispensada al paciente y uno relativo a la identificación del mismo. Paralelamente, se diseñó un protocolo de trabajo estandarizado y se evaluó el cumplimiento de los criterios sobre todos los cajetines de medicación de los pacientes hospitalizados en dos unidades clínicas. El cumplimiento de los criterios se midió antes y después de la implantación del protocolo, durante catorce y siete días consecutivos respectivamente. Resultados: En la primera evaluación el número de incumplimientos fue 0,84/cajetín. Tras la implantación del protocolo, el total de incumplimientos tras ajuste por tamaño de muestra se redujo a 0,22/cajetín. El nivel de cumplimiento mejoró para todos los criterios y las diferencias fueron significativas para los dos criterios más incumplidos. Conclusiones: La implantación de un protocolo ha supuesto una mejora significativa en dos de los cinco criterios estudiados en el proceso llenado de carros de medicación. La identificación del paciente se ha revelado como un aspecto fundamental de intervención en la calidad de la dispensación.