Sugar metabolism, redox balance and oxidative stress response in the respiratory yeast Kluyveromyces lactis

A lot of studies have been carried out on Saccharomyces cerevisiae, an yeast with a predominant fermentative metabolism under aerobic conditions, which allows exploring the complex response induced by oxidative stress. S. cerevisiae is considered a eukaryote model for these studies. We propose Kluyveromyces lactis as a good alternative model to analyse variants in the oxidative stress response, since the respiratory metabolism in this yeast is predominant under aerobic conditions and it shows other important differences with S. cerevisiae in catabolic repression and carbohydrate utilization. The knowledge of oxidative stress response in K. lactis is still a developing field. In this article, we summarize the state of the art derived from experimental approaches and we provide a global vision on the characteristics of the putative K. lactis components of the oxidative stress response pathway, inferred from their sequence homology with the S. cerevisiae counterparts. Since K. lactis is also a well-established alternative host for industrial production of native enzymes and heterologous proteins, relevant differences in the oxidative stress response pathway and their potential in biotechnological uses of this yeast are also reviewed

Enhancement of the bioproduction potential of an amylaceous effluent

9 páginas, 12 figuras, 4 tablasThe treatment of starchy effluents could provide the basis for a series of simultaneous or alternative bioproductions easily integrable into a system of greater economic attraction that the simple production of SCP. In order to define some of the possible options, this study uses an amylaceous effluent for simultaneous production of SCP and an amylolytic preparation which is partially consumed in the saccharification of a parallel flow of the same effluent concentrated by ultrafiltration with cutoff at 100 kD. With regard to the development of the above system, this paper describes the problems associated with ultrafiltration, the conditions that optimize saccharification of the concentrate and the requirements of various possible bioproductions that couM be obtained from the saccharified concentrate.CICYT (project ALl 789- 9O).Peer reviewe

Biochemical and structural characterization of a novel thermophilic esterase EstD11 provide catalytic insights for the HSL family

19 pags., 12 figs., 2 tabs.A novel esterase, EstD11, has been discovered in a hot spring metagenomic library. It is a thermophilic and thermostable esterase with an optimum temperature of 60°C. A detailed substrate preference analysis of EstD11 was done using a library of chromogenic ester substrate that revealed the broad substrate specificity of EstD11 with significant measurable activity against 16 substrates with varied chain length, steric hindrance, aromaticity and flexibility of the linker between the carboxyl and the alcohol moiety of the ester. The tridimensional structures of EstD11 and the inactive mutant have been determined at atomic resolutions. Structural and bioinformatic analysis, confirm that EstD11 belongs to the family IV, the hormone-sensitive lipase (HSL) family, from the α/β-hydrolase superfamily. The canonical α/β-hydrolase domain is completed by a cap domain, composed by two subdomains that can unmask of the active site to allow the substrate to enter. Eight crystallographic complexes were solved with different substrates and reaction products that allowed identification of the hot-spots in the active site underlying the specificity of the protein. Crystallization and/or incubation of EstD11 at high temperature provided unique information on cap dynamics and a first glimpse of enzymatic activity in vivo. Very interestingly, we have discovered a unique Met zipper lining the active site and the cap domains that could be essential in pivotal aspects as thermo-stability and substrate promiscuity in EstD11.We thank the staff from ALBA synchrotron facility (Barcelona, Spain) for support during crystallographic data collection. This work was funded by the UE through the HotDrops Project (FP7- PEOPLE-2012-IAPP, project number 324439). Additionally, this work was supported by the BFU2017-90030-P grant to J.A.H. from the Spanish Ministry of Science and Innovatio

Genome-Wide Analysis of the Yeast Transcriptome Upon Heat and Cold Shock

DNA arrays were used to measure changes in transcript levels as yeast cells responded to temperature shocks. The number of genes upregulated by temperature shifts from 30 ℃ to 37℃ or 45℃ was correlated with the severity of the stress. Pre-adaptation of cells, by growth at 37 ℃ previous to the 45℃ shift, caused a decrease in the number of genes related to this response. Heat shock also caused downregulation of a set of genes related to metabolism, cell growth and division, transcription, ribosomal proteins, protein synthesis and destination. Probably all of these responses combine to slow down cell growth and division during heat shock, thus saving energy for cell rescue. The presence of putative binding sites for Xbp1p in the promoters of these genes suggests a hypothetical role for this transcriptional repressor, although other mechanisms may be considered. The response to cold shock (4℃) affected a small number of genes, but the vast majority of those genes induced by exposure to 4 ℃ were also induced during heat shock; these genes share in their promoters cis-regulatory elements previously related to other stress responses

Biochemical and structural characterization of a novel thermophilic esterase EstD11 provide catalytic insights for the HSL family

[Abstract]: A novel esterase, EstD11, has been discovered in a hot spring metagenomic library. It is a thermophilic and thermostable esterase with an optimum temperature of 60 C. A detailed substrate preference analysis of EstD11 was done using a library of chromogenic ester substrate that revealed the broad substrate specificity of EstD11 with significant measurable activity against 16 substrates with varied chain length, steric hindrance, aromaticity and flexibility of the linker between the carboxyl and the alcohol moiety of the ester. The tridimensional structures of EstD11 and the inactive mutant have been determined at atomic resolutions. Structural and bioinformatic analysis, confirm that EstD11 belongs to the family IV, the hormone-sensitive lipase (HSL) family, from the α/β-hydrolase superfamily. The canonical a/b hydrolase domain is completed by a cap domain, composed by two subdomains that can unmask of the active site to allow the substrate to enter. Eight crystallographic complexes were solved with different substrates and reaction products that allowed identification of the hot-spots in the active site underlying the specificity of the protein. Crystallization and/or incubation of EstD11 at high temperature provided unique information on cap dynamics and a first glimpse of enzymatic activity in vivo. Very interestingly, we have discovered a unique Met zipper lining the active site and the cap domains that could be essential in pivotal aspects as thermo-stability and substrate promiscuity in EstD11Ministerio de Ciencia e Innovación; BFU2017-90030-

A motivational interview program for cardiac rehabilitation after acute myocardial infarction: study protocol of a randomized controlled trial in primary healthcare

Cardiac rehabilitation after acute myocardial infarction permits recovery of the heart function and enables secondary prevention programs in which changes in lifestyle habits are crucial. Cardiac rehabilitation often takes place in hospitals without coordination with primary healthcare and is not focused on individual patient preferences and goals, which is the core of the motivational interview. The objective of this study was to evaluate the efficacy of a cardiac rehabilitation program with a motivational interview in patients discharged from hospital after acute myocardial infarction.A randomized, non-pharmacological clinical trial in six primary healthcare centers in Barcelona (Spain) will assess whether a tailored cardiac rehabilitation program consisting of four motivational interviews and visits with family physicians, primary healthcare nurses and a cardiologist, coordinated with the reference hospital, results in better cardiac rehabilitation than standard care. A minimum sample of 284 participants requiring cardiac rehabilitation after acute myocardial infarction will be randomized to a cardiac rehabilitation group with a motivational interview program or to standard primary healthcare. The main outcome will be physical function measured by the six-minute walk test, and the secondary outcome will be the effectiveness of secondary prevention: a composite outcome comprising control of blood pressure, cholesterol, diabetes mellitus, smoking and body weight. Results will be evaluated at 1,3 and 6 months.This is the first clinical trial to study the impact of a new primary healthcare cardiac rehabilitation program with motivational interviews for patients discharged from hospital after myocardial infarction. Changes in lifestyles and habits after myocardial infarction are a core element of secondary prevention and require patient-centered care strategies such as motivational interviews. Therefore, this study could clarify the impact of this approach on health indicators, such as functional capacity.ClinicalTriasl.gov NCT05285969 registered on March 18, 2022.© 2022. The Author(s)

The impact of oxygen on the transcriptome of recombinant S. cerevisiae and P. pastoris - a comparative analysis

Background: Saccharomyces cerevisiae and Pichia pastoris are two of the most relevant microbial eukaryotic platforms for the production of recombinant proteins. Their known genome sequences enabled several transcriptomic profiling studies under many different environmental conditions, thus mimicking not only perturbations and adaptations which occur in their natural surroundings, but also in industrial processes. Notably, the majority of such transcriptome analyses were performed using non-engineered strains. In this comparative study, the gene expression profiles of S. cerevisiae and P. pastoris, a Crabtree positive and Crabtree negative yeast, respectively, were analyzed for three different oxygenation conditions (normoxic, oxygen-limited and hypoxic) under recombinant protein producing conditions in chemostat cultivations. Results: The major differences in the transcriptomes of S. cerevisiae and P. pastoris were observed between hypoxic and normoxic conditions, where the availability of oxygen strongly affected ergosterol biosynthesis, central carbon metabolism and stress responses, particularly the unfolded protein response. Steady state conditions under low oxygen set-points seemed to perturb the transcriptome of S. cerevisiae to a much lesser extent than the one of P. pastoris, reflecting the major tolerance of the baker's yeast towards oxygen limitation, and a higher fermentative capacity. Further important differences were related to Fab production, which was not significantly affected by oxygen availability in S. cerevisiae, while a clear productivity increase had been previously reported for hypoxically grown P. pastoris. Conclusions: The effect of three different levels of oxygen availability on the physiology of P. pastoris and S. cerevisiae revealed a very distinct remodelling of the transcriptional program, leading to novel insights into the different adaptive responses of Crabtree negative and positive yeasts to oxygen availability. Moreover, the application of such comparative genomic studies to recombinant hosts grown in different environments might lead to the identification of key factors for efficient protein production