Testing Biochemistry Revisited: How In Vivo Metabolism Can Be Understood from In Vitro Enzyme Kinetics

A decade ago, a team of biochemists including two of us, modeled yeast glycolysis and showed that one of the most studied biochemical pathways could not be quite understood in terms of the kinetic properties of the constituent enzymes as measured in cell extract. Moreover, when the same model was later applied to different experimental steady-state conditions, it often exhibited unrestrained metabolite accumulation

Return-to-work of sick-listed workers without an employment contract – what works?

BACKGROUND: In the past decade flexible labour market arrangements have emerged as a significant change in the European Union labour market. Studies suggest that these new types of labour arrangements may be linked to ill health, an increased risk for work disability, and inadequate vocational rehabilitation. Therefore, the objectives of this study were: 1. to examine demographic characteristics of workers without an employment contract sick-listed for at least 13 weeks, 2. to describe the content and frequency of occupational health care (OHC) interventions for these sick-listed workers, and 3. to examine OHC interventions as possible determinants for return-to-work (RTW) of these workers. METHODS: A cohort of 1077 sick-listed workers without an employment contract were included at baseline, i.e. 13 weeks after reporting sick. Demographic variables were available at baseline. Measurement of cross-sectional data took place 4-6 months after inclusion. Primary outcome measures were: frequency of OHC interventions and RTW-rates. Measured confounding variables were: gender, age, type of worker (temporary agency worker, unemployed worker, or remaining worker without employment contract), level of education, reason for absenteeism (diagnosis), and perceived health. The association between OHC interventions and RTW was analysed with a logistic multiple regression analysis. RESULTS: At 7-9 months after the first day of reporting sick only 19% of the workers had (partially or completely) returned to work, and most workers perceived their health as fairly poor or poor. The most frequently reported (49%) intervention was 'the OHC professional discussed RTW'. However, the intervention 'OHC professional made and discussed a RTW action plan' was reported by only 19% of the respondents. The logistic multiple regression analysis showed a significant positive association between RTW and the interventions: 'OHC professional discussed RTW'; and 'OHC professional made and discussed a RTW action plan'. The intervention 'OHC professional referred sick-listed worker to a vocational rehabilitation agency' was significantly associated with no RTW. CONCLUSION: This is the first time that characteristics of a large cohort of sick-listed workers without an employment contract were examined. An experimental or prospective study is needed to explore the causal nature of the associations found between OHC interventions and RT

Evidence for Divergent Evolution of Growth Temperature Preference in Sympatric Saccharomyces Species

The genus Saccharomyces currently includes eight species in addition to the model yeast Saccharomyces cerevisiae, most of which can be consistently isolated from tree bark and soil. We recently found sympatric pairs of Saccharomyces species, composed of one cryotolerant and one thermotolerant species in oak bark samples of various geographic origins. In order to contribute to explain the occurrence in sympatry of Saccharomyces species, we screened Saccharomyces genomic data for protein divergence that might be correlated to distinct growth temperature preferences of the species, using the dN/dS ratio as a measure of protein evolution rates and pair-wise species comparisons. In addition to proteins previously implicated in growth at suboptimal temperatures, we found that glycolytic enzymes were among the proteins exhibiting higher than expected divergence when one cryotolerant and one thermotolerant species are compared. By measuring glycolytic fluxes and glycolytic enzymatic activities in different species and at different temperatures, we subsequently show that the unusual divergence of glycolytic genes may be related to divergent evolution of the glycolytic pathway aligning its performance to the growth temperature profiles of the different species. In general, our results support the view that growth temperature preference is a trait that may have undergone divergent selection in the course of ecological speciation in Saccharomyces

Mathematical modeling of intracellular signaling pathways

Dynamic modeling and simulation of signal transduction pathways is an important topic in systems biology and is obtaining growing attention from researchers with experimental or theoretical background. Here we review attempts to analyze and model specific signaling systems. We review the structure of recurrent building blocks of signaling pathways and their integration into more comprehensive models, which enables the understanding of complex cellular processes. The variety of mechanisms found and modeling techniques used are illustrated with models of different signaling pathways. Focusing on the close interplay between experimental investigation of pathways and the mathematical representations of cellular dynamics, we discuss challenges and perspectives that emerge in studies of signaling systems

Capillary electrophoresis mass spectrometry-based saliva metabolomics identified oral, breast and pancreatic cancer-specific profiles

Saliva is a readily accessible and informative biofluid, making it ideal for the early detection of a wide range of diseases including cardiovascular, renal, and autoimmune diseases, viral and bacterial infections and, importantly, cancers. Saliva-based diagnostics, particularly those based on metabolomics technology, are emerging and offer a promising clinical strategy, characterizing the association between salivary analytes and a particular disease. Here, we conducted a comprehensive metabolite analysis of saliva samples obtained from 215 individuals (69 oral, 18 pancreatic and 30 breast cancer patients, 11 periodontal disease patients and 87 healthy controls) using capillary electrophoresis time-of-flight mass spectrometry (CE-TOF-MS). We identified 57 principal metabolites that can be used to accurately predict the probability of being affected by each individual disease. Although small but significant correlations were found between the known patient characteristics and the quantified metabolites, the profiles manifested relatively higher concentrations of most of the metabolites detected in all three cancers in comparison with those in people with periodontal disease and control subjects. This suggests that cancer-specific signatures are embedded in saliva metabolites. Multiple logistic regression models yielded high area under the receiver-operating characteristic curves (AUCs) to discriminate healthy controls from each disease. The AUCs were 0.865 for oral cancer, 0.973 for breast cancer, 0.993 for pancreatic cancer, and 0.969 for periodontal diseases. The accuracy of the models was also high, with cross-validation AUCs of 0.810, 0.881, 0.994, and 0.954, respectively. Quantitative information for these 57 metabolites and their combinations enable us to predict disease susceptibility. These metabolites are promising biomarkers for medical screening

Can yeast glycolysis be understood in terms of in vitro kinetics of the constituent enxymes? Testing biochemistry.

This paper examines whether the in vivo behavior of yeast glycolysis can be understood in terms of the in vitro kinetic properties of the constituent enzymes. In nongrowing, anaerobic, compressed Saccharomyces cerevisiae the values of the kinetic parameters of most glycolytic enzymes were determined. For the other enzymes appropriate literature values were collected. By inserting these values into a kinetic model for glycolysis, fluxes and metabolites were calculated. Under the same conditions fluxes and metabolite levels were measured. In our first model, branch reactions were ignored. This model failed to reach the stable steady state that was observed in the experimental flux measurements. Introduction of branches towards trehalose, glycogen, glycerol and succinate did allow such a steady state. The predictions of this branched model were compared with the empirical behavior. Half of the enzymes matched their predicted flux in vivo within a factor of 2. For the other enzymes it was calculated what deviation between in vivo and in vitro kinetic characteristics could explain the discrepancy between in vitro rate and in vivo flux

Can yeast glycolysis be understood in terms of <I>in vitro</I> kinetics of the constituent enzymes?

NatuurwetenskappeBiochemiePlease help us populate SUNScholar with the post print version of this article. It can be e-mailed to: [email protected]