Related ArticlesSpectral Comparisons of Mammalian Cells and Intact Organelles by Solid-State NMR.
J Struct Biol. 2018 May 30;:
Authors: Werby SH, Cegelski L
Abstract
Whole-cell protein profiling, spatial localization, and quantification of activities such as gene transcription and protein translation are possible with modern biochemical and biophysical techniques. Yet, addressing questions of overall compositional changes within a cell - capturing the relative amounts of protein and ribosomal RNA levels and lipid content simultaneously - would require numerous extractions and purifications with caveats due to isolation yields and detection methods. A holistic view of cellular composition would aid in the study of cellular composition and function. Here, solid state NMR is used to identify 13C NMR signatures for cellular organelles in HeLa cells, without the use of any isotopic labeling. Comparisons are made with carbon spectra of subcellular assemblies including DNA, lipids, ribosomes, nuclei and mitochondria. Whole-cell comparisons are made with different mammalian cells lines, with red blood cells that lack nuclei and organelles, and with Gram-negative and Gram-positive bacteria. Furthermore, treatment of mammalian cells with cycloheximide, a commonly used protein synthesis inhibitor, revealed unanticipated changes consistent with a significant increase in protein glycosylation, obvious at the whole cell level. Thus, we demonstrate that solid-state NMR serves as a unique analytical tool to catalog and compare the ratios of distinct carbon types in cells and serves as a discovery tool to reveal the workings of inhibitors such as cycloheximide on whole-cell biochemistry.
PMID: 29859329 [PubMed - as supplied by publisher]
[NMR paper] Preparation of highly and generally enriched mammalian tissues for solid state NMR.
Preparation of highly and generally enriched mammalian tissues for solid state NMR.
http://www.bionmr.com//www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--production.springer.de-OnlineResources-Logos-springerlink.gif Related Articles Preparation of highly and generally enriched mammalian tissues for solid state NMR.
J Biomol NMR. 2015 Oct;63(2):119-23
Authors: Wong VW, Reid DG, Chow WY, Rajan R, Green M, Brooks RA, Duer MJ
Abstract
An appreciable level of isotope labelling is essential for future NMR structure elucidation of...
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07-02-2016 07:23 PM
[NMR paper] A Genetically Encoded ?-Lactamase Reporter for Ultrasensitive (129) Xe NMR in Mammalian Cells.
A Genetically Encoded ?-Lactamase Reporter for Ultrasensitive (129) Xe NMR in Mammalian Cells.
Related Articles A Genetically Encoded ?-Lactamase Reporter for Ultrasensitive (129) Xe NMR in Mammalian Cells.
Angew Chem Int Ed Engl. 2016 Jun 15;
Authors: Wang Y, Roose BW, Palovcak EJ, Carnevale V, Dmochowski IJ
Abstract
Molecular imaging holds considerable promise for elucidating biological processes in normal physiology as well as disease states, but requires noninvasive methods for identifying analytes at sub-micromolar...
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06-16-2016 12:06 PM
[NMR paper] Characterization of proteins by in-cell NMR spectroscopy in cultured mammalian cells.
Characterization of proteins by in-cell NMR spectroscopy in cultured mammalian cells.
Characterization of proteins by in-cell NMR spectroscopy in cultured mammalian cells.
Nat Protoc. 2016 Jun;11(6):1101-1111
Authors: Barbieri L, Luchinat E, Banci L
Abstract
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05-20-2016 03:04 PM
[NMR paper] Algal autolysate medium to label proteins for NMR in mammalian cells.
Algal autolysate medium to label proteins for NMR in mammalian cells.
Related Articles Algal autolysate medium to label proteins for NMR in mammalian cells.
J Biomol NMR. 2016 Apr 22;
Authors: Fuccio C, Luchinat E, Barbieri L, Neri S, Fragai M
Abstract
In-cell NMR provides structural and functional information on proteins directly inside living cells. At present, the high costs of the labeled media for mammalian cells represent a limiting factor for the development of this methodology. Here we report a protocol to prepare a...
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04-24-2016 10:29 PM
Algal autolysate medium to label proteins for NMR in mammalian cells
Algal autolysate medium to label proteins for NMR in mammalian cells
Abstract
In-cell NMR provides structural and functional information on proteins directly inside living cells. At present, the high costs of the labeled media for mammalian cells represent a limiting factor for the development of this methodology. Here we report a protocol to prepare a homemade growth medium from Spirulina platensis autolysate, suitable to express uniformly labeled proteins inside mammalian cells at a reduced cost-per-sample. The human proteins SOD1 and Mia40 were...
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04-22-2016 08:45 PM
[NMR paper] Fluorine NMR-based assay in living mammalian cells.
Fluorine NMR-based assay in living mammalian cells.
http://www.bionmr.com//www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--linkinghub.elsevier.com-ihub-images-PubMedLink.gif Related Articles Fluorine NMR-based assay in living mammalian cells.
Anal Biochem. 2015 Dec 11;
Authors: Veronesi M, Giacomina F, Romeo E, Castellani B, Ottonello G, Lambruschini C, Garau G, Scarpelli R, Bandiera T, Piomelli D, Dalvit C
Abstract
Nuclear Magnetic Resonance (NMR)-based screening has been recognized as a powerful approach for the identification...
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12-28-2015 12:26 AM
Preparation of highly and generally enriched mammalian tissues for solid state NMR
Preparation of highly and generally enriched mammalian tissues for solid state NMR
Abstract
An appreciable level of isotope labelling is essential for future NMR structure elucidation of mammalian biomaterials, which are either poorly expressed, or unexpressable, using micro-organisms. We present a detailed protocol for high level 13C enrichment even in slow turnover murine biomaterials (fur keratin), using a customized diet supplemented with commercial labelled algal hydrolysate and formulated as a gel to minimize wastage, which female mice...
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09-26-2015 05:13 AM
[NMR paper] Detection of fructose-3-phosphokinase activity in intact mammalian lenses by 31P NMR
Detection of fructose-3-phosphokinase activity in intact mammalian lenses by 31P NMR spectroscopy.
Related Articles Detection of fructose-3-phosphokinase activity in intact mammalian lenses by 31P NMR spectroscopy.
J Biol Chem. 1993 Apr 15;268(11):7763-7
Authors: Lal S, Szwergold BS, Kappler F, Brown T
Recently we have identified two novel phosphorylated metabolites in the lenses of diabetic rats as sorbitol 3-phosphate (Sor-3-P) and fructose 3-phosphate (Fru-3-P). The latter compound is of particular interest since it is a potent glycating...