<i>In Situ</i>NMR Systems

Shanks, Jacqueline V.

doi:10.21775/cimb.003.015

Current Issues in Molecular Biology

2001

DOI: 10.21775/cimb.003.015

|View full text |Cite

In SituNMR Systems

Jacqueline V. Shanks

Abstract: In situ NMR is becoming an established technology for applications in bioprocessing and metabolic engineering. The in situ NMR biosensor acts as a noninvasive pH, ion, and concentration meter, with 31 P and 13 C as the two main isotopes of study. A substantial data base now exists for phosphorus and carbon spectra of bacteria and yeast. In situ NMR can provide many of the state variables needed for modeling glycolytic pathway function. NMR micro-reactor technology has improved significantly in the last decade.… Show more

Help me understand this report

Search citation statements

Order By: Relevance

Paper Sections

Select...

Citation Types

Supporting

Mentioning

Contrasting

Year Published

2002

2022

Publication Types

Select...

Article4

Relationship

Self Cite0

Independent4

Authors

Journals

Cited by 4 publications

References 56 publications

(145 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

Untitled

Ramos

Neves

Santos

2002

Antonie Van Leeuwenhoek

View full text Add to dashboard Cite

The complexity of metabolic and regulatory networks presents a great scientific challenge to an integrated view of how individual components contribute to the overall function. Nuclear magnetic resonance (NMR) spectroscopy is undoubtedly a suitable technique for global investigations of microbial metabolism, since it allows a view into living cells without disturbing the cellular organisation. Therefore, metabolic processes can be monitored in real time under physiological conditions. In the present paper, examples of the application of NMR to study the metabolism of lactic acid bacteria will be given. These include the analysis of labelling patterns in end-products using 13C as a tracer, thereby establishing metabolic pathways, the detection and quantification of intermediates in the pathway of exopolysaccharide biosynthesis, and on line monitoring of glycolytic kinetics to assess the effect of metabolic engineering strategies.

show abstract

Untitled

Ramos

Neves

Santos

2002

Antonie Van Leeuwenhoek

View full text Add to dashboard Cite

show abstract

The Energetic Conversion Competence of Escherichia coli during Aerobic Respiration Studied by 31P NMR Using a Circulating Fermentation System

Noguchi¹

2004

Journal of Biochemistry

View full text Add to dashboard Cite

To determine the actual potential of the energetic conversion efficiency of Escherichia coli during aerobic respiration, apparent P/O ratios (P/O(app)) under either limited or standard glucose-feeding conditions were estimated. The previously reported circulating fermentation system (CFS) was used, and (31)P NMR saturation-transfer (ST) techniques were employed. By coupling with on-line NMR observations, CFS allowed us to evaluate cellular energetics directly, with both the dissolved oxygen tension and glucose feeding precisely controlled to prevent the effect of substrate-level phosphorylation based on aerobic or anaerobic acidogenesis in E. coli cells. Phosphate consumption rates under standard and limited glucose-conditions were estimated as 4.62 +/- 0.46 and 1.99 +/- 0.11 micromol/s g of dry cell weight (DCW), respectively. Using simultaneously assessed O(2) consumption rates, the P/O(app) values under these two conditions were estimated as 1.4 +/- 0.3 and 1.5 +/- 0.1, respectively. To correlate the obtained P/O(app) values with the potential efficiency of respiratory enzymes, we determined the activities of two NADH dehydrogenases (NDH 1 and 2) and two ubiquinol oxidases (bo- and bd-type) during the periods when ST was performed. NDH-1 activities in standard or limited glucose cultures were maintained at 57% or 58% of the total NADH oxidizing activity. The percentages of bo-type oxidase activity in relation to the total ubiqinol oxidizing activity under the standard and limited glucose conditions were 32% and 36%, respectively. These percentages of enzymatic activities represent the respiratory competence of E. coli cells, suggesting that, during the NMR observatory period, the enzymatic activity was not at a maximum, which could also explain the estimated P/O(app) values. If this is the case, enhancing the expression of the bo-type oxidase or disrupting of the bd-type oxidase gene could be effective approach to increasing both the P/O ratio and cellular yields.

show abstract

Targeted redox and energy cofactor metabolomics in Clostridium thermocellum and Thermoanaerobacterium saccharolyticum

Sander

Asano

Bhandari

et al. 2017

Biotechnol Biofuels

View full text Add to dashboard Cite

Background Clostridium thermocellum and Thermoanaerobacterium saccharolyticum are prominent candidate biocatalysts that, together, can enable the direct biotic conversion of lignocellulosic biomass to ethanol. The imbalance and suboptimal turnover rates of redox cofactors are currently hindering engineering efforts to achieve higher bioproductivity in both organisms. Measuring relevant intracellular cofactor concentrations will help understand redox state of these cofactors and help identify a strategy to overcome these limitations; however, metabolomic determinations of these labile metabolites have historically proved challenging.ResultsThrough our validations, we verified the handling and storage stability of these metabolites, and verified extraction matrices and extraction solvent were not suppressing mass spectrometry signals. We recovered adenylate energy charge ratios (a main quality indicator) above 0.82 for all extractions. NADH/NAD+ values of 0.26 and 0.04 for an adhE-deficient strain of C. thermocellum and its parent, respectively, reflect the expected shift to a more reduced redox potential when a species lacks the ability to re-oxidize NADH by synthesizing ethanol. This method failed to yield reliable results with C. bescii and poor-growing strains of T. saccharolyticum. ConclusionsOur validated protocols demonstrate and validate the extraction and analysis of selected redox and energy-related metabolites from two candidate consolidated bioprocessing biocatalysts, C. thermocellum and T. saccharolyticum. This development and validation highlights the important, but often neglected, need to optimize and validate metabolomic protocols when adapting them to new cell or tissue types.

show abstract

scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.

Contact Info

customersupport@researchsolutions.com

10624 S. Eastern Ave., Ste. A-614

Henderson, NV 89052, USA

This site is protected by reCAPTCHA and the Google Privacy Policy and Terms of Service apply.

Blog Terms and Conditions API Terms Privacy Policy Contact Cookie Preferences Do Not Sell or Share My Personal Information

Made with 💙 for researchers

Part of the Research Solutions Family.

In SituNMR Systems

Cited by 4 publications

References 56 publications

Untitled

Untitled

The Energetic Conversion Competence of Escherichia coli during Aerobic Respiration Studied by 31P NMR Using a Circulating Fermentation System

Targeted redox and energy cofactor metabolomics in Clostridium thermocellum and Thermoanaerobacterium saccharolyticum

Contact Info

Product

Resources

About