Semi-on-line analysis for fast and precise monitoring of bioreaction processes

Christiansen, Lasse Engbo; Marcher, J.; Schulze, Ulrik; Carlsen, Morten; Min, Rong; Nielsen, Jens

doi:10.1002/(sici)1097-0290(19961020)52:2<237::aid-bit5>3.0.co;2-r

Cited by 24 publications

(14 citation statements)

References 43 publications

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“…The temperature was kept constant at 308C and pH was maintained at 5.00 AE 0.05 by addition of KOH. The concentration of carbon dioxide and oxygen in the exhaust gas was determined by use of a Brüel and Kjaer acoustic gas analyzer (Brüel & Kjaer, Naerum, Denmark) (Christensen et al, 1995). We carried out biological independent triplicate cultures for each mutant.…”

Section: Batch Cultivationsmentioning

confidence: 99%

A systems biology approach to study glucose repression in the yeast Saccharomyces cerevisiae

Westergaard

Oliveira

Bro

et al. 2006

Biotech & Bioengineering

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Glucose repression in the yeast Saccharomyces cerevisiae has evolved as a complex regulatory system involving several different pathways. There are two main pathways involved in signal transduction. One has a role in glucose sensing and regulation of glucose transport, while another takes part in repression of a wide range of genes involved in utilization of alternative carbon sources. In this work, we applied a systems biology approach to study the interaction between these two pathways. Through genome-wide transcription analysis of strains with disruption of HXK2, GRR1, MIG1, the combination of MIG1 and MIG2, and the parental strain, we identified 393 genes to have significantly changed expression levels. To identify co-regulation patterns in the different strains we applied principal component analysis. Disruption of either GRR1 or HXK2 were both found to have profound effects on transcription of genes related to TCA cycle and respiration, as well as ATP synthesis coupled proton transport, all displaying an increased expression. The hxk2Delta strain showed reduced overflow metabolism towards ethanol relative to the parental strain. We also used a genome-scale metabolic model to identify reporter metabolites, and found that there is a high degree of consistency between the identified reporter metabolites and the physiological effects observed in the different mutants. Our systems biology approach points to close interaction between the two pathways, and our metabolism driven analysis of transcription data may find a wider application for analysis of cross-talk between different pathways involved in regulation of metabolism.

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Section: Batch Cultivationsmentioning

confidence: 99%

A systems biology approach to study glucose repression in the yeast Saccharomyces cerevisiae

Westergaard

Oliveira

Bro

et al. 2006

Biotech & Bioengineering

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“…The ability to monitor the concentrations of nutrients and byproducts in cell culture bioreactors is necessary for the development of intelligent bioreactor control schemes. Flow injection systems and off‐line analyses provide accurate evaluation of the medium composition, however, they also require that samples be removed from the reactor prior to analysis (Christensen et al, 1996; Ozturk et al, 1997). This removal step can create concerns for bioreactor contamination, can restrict the number of samples taken from a small bioreactor, and can complicate automation schemes.…”

Section: Introductionmentioning

confidence: 99%

Adaptive Calibration Scheme for Quantification of Nutrients and Byproducts in Insect Cell Bioreactors by Near-Infrared Spectroscopy

et al. 1998

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Spectroscopic methods are gaining in popularity in biotechnology because of their ability to deliver rapid, noninvasive measurements of the concentrations of multiple chemical species. Such measurements are particularly necessary for the implementation of control schemes for cell culture bioreactors. One of the major challenges to the development of spectroscopic methods for bioreactor monitoring is the generation of accurate and robust calibration models, particularly because of the inherent variability of biological processes. We have evaluated several methods of building calibration models, including synthetic calibrations and medium spiking methods. The approach that consistently produced reliable models incorporated samples removed from a bioreactor that were subsequently altered so as to increase the sample variation. Several large volume samples were removed from a bioreactor at varying time points and divided into multiple aliquots to which were added random, known amounts of the analytes of interest. Near-infrared spectra of these samples were collected and used to build calibration models. Such models were used to quantify analyte concentrations from independent samples removed from a second bioreactor. Prediction errors for alanine, glucose, glutamine, and leucine were 1.4, 1.0, 1.1, and 0.31 mM, respectively. This adaptive calibration method produces models with less error and less bias than observed with other calibration methods. Somewhat more accurate measurements could be attained with calibrations consisting of a combination of synthetic samples and spiked medium samples, but with an increase in calibration development time.

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“…Carbon dioxide and oxygen concentrations in the exhaust gas were determined by a Brüel and Kjaer (Naerum, Denmark) 1308 acoustic gas analyzer (2).…”

Section: Methodsmentioning

confidence: 99%

Metabolic Engineering of Ammonium Assimilation in Xylose-Fermenting Saccharomyces cerevisiae Improves Ethanol Production

Roca

Nielsen

Olsson

2003

Appl Environ Microbiol

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Cofactor imbalance impedes xylose assimilation in Saccharomyces cerevisiae that has been metabolically engineered for xylose utilization. To improve cofactor use, we modified ammonia assimilation in recombinant S. cerevisiae by deleting GDH1, which encodes an NADPH-dependent glutamate dehydrogenase, and by overexpressing either GDH2, which encodes an NADH-dependent glutamate dehydrogenase, or GLT1 and GLN1, which encode the GS-GOGAT complex. Overexpression of GDH2 increased ethanol yield from 0.43 to 0.51 mol of carbon (Cmol) Cmol ؊1 , mainly by reducing xylitol excretion by 44%. Overexpression of the GS-GOGAT complex did not improve conversion of xylose to ethanol during batch cultivation, but it increased ethanol yield by 16% in carbon-limited continuous cultivation at a low dilution rate.In order to develop an efficient process for the production of bioethanol from lignocellulosic material, there have been many attempts to improve the conversion of xylose to ethanol by construction of recombinant Saccharomyces cerevisiae strains.Even though the open reading frames encoding the three first enzymes involved in xylose metabolism, i.e., xylose reductase (XR), xylitol dehydrogenase (XDH), and xylulose kinase (XK), have been identified in the genome of S. cerevisiae (10, 15), the genes are poorly expressed (or not expressed at all), and xylose cannot be metabolized without introduction of heterologous genes from naturally xylose-fermenting microorganisms and overexpression of endogenous XK (3,6,17). In the resulting strains, there is a low rate of xylose consumption and substantial xylitol secretion.Since xylose-metabolizing strains of S. cerevisiae harbor a NAD(P)H-dependent xylose reductase and a NAD ϩ -dependent xylitol dehydrogenase from Pichia stipitis, one of the main reasons for xylitol excretion can be explained by the redox imbalance occurring at the level of XR and XDH. Metabolic engineering of redox metabolism may therefore be a strategy for improving the conversion of xylose to ethanol. This has been illustrated by deletion of the zwf1 gene encoding glucose-6-phosphate dehydrogenase in a xylose-metabolizing strain of S. cerevisiae (7). The main source of NADPH originating from the oxidative part of the pentose phosphate pathway has thereby been reduced, and during growth on xylose, this has resulted in a significant improvement of ethanol yield, from 0.31 g g Ϫ1 (yield on consumed sugar) to 0.41 g g Ϫ1 with a concomitant reduction in the xylitol yield.In this study, we have taken a different approach to modulating the redox metabolism to favor xylose metabolism, namely through metabolic engineering of the ammonia assimilation. Ammonium assimilation in S. cerevisiae involves glutamate dehydrogenase and glutamine synthetase. Glutamate dehydrogenase catalyzes the synthesis of glutamate from ammonium and 2-ketoglutarate. The most important enzyme for ammonia assimilation is the NADPH-dependent glutamate dehydrogenase 1, encoded by GDH1. The other glutamate dehydrogenase that is present in S. cerevisiae is N...

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Semi-on-line analysis for fast and precise monitoring of bioreaction processes

Cited by 24 publications

References 43 publications

A systems biology approach to study glucose repression in the yeast Saccharomyces cerevisiae

A systems biology approach to study glucose repression in the yeast Saccharomyces cerevisiae

Adaptive Calibration Scheme for Quantification of Nutrients and Byproducts in Insect Cell Bioreactors by Near-Infrared Spectroscopy

Metabolic Engineering of Ammonium Assimilation in Xylose-Fermenting Saccharomyces cerevisiae Improves Ethanol Production

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