Optimization of Metabolic Labeling with Alkyne-Containing Isoprenoid Probes

The concentration of glycogen, the major brain energy reserve localized mainly in astrocytes, is generally reported as about 2 or 3 μmol/g, but sometimes as high as 3.9 to 8 μmol/g, in normal rat brain. The authors found high but very different glycogen levels in two recent studies in which glycogen was determined by the routine amyloglucosidase procedure in 0.03N HCl digests either of frozen powders (4.8 to 6 μmol/g) or of ethanol-insoluble fractions (8 to 12 μmol/g). To evaluate the basis for these discrepant results, glycogen was assayed in parallel extracts of the same samples. Glycogen levels in ethanol extracts were twice those in 0.03N HCl digests, suggesting incomplete enzyme inactivation even with very careful thawing. The very high glycogen levels were biologically active and responsive to physiologic and pharmacological challenge. Glycogen levels fell after brief sensory stimulation, and metabolic labeling indicated its turnover under resting conditions. About 95% of the glycogen was degraded under in vitro ischemic conditions, and its “carbon equivalents” recovered mainly as glc, glc-P, and lactate. Resting glycogen stores were reduced by about 50% by chronic inhibition of nitric oxide synthase. Because neurotransmitters are known to stimulate glycogenolysis, stress or sensory activation due to animal handling and tissue-sampling procedures may stimulate glycogenolysis during an experiment, and glycogen lability during tissue sampling and extraction can further reduce glycogen levels. The very high glycogen levels in normal rat brain suggest an unrecognized role for astrocytic energy metabolism during brain activation.

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Using Heavy Mass Isotopomers for Protein Turnover in Heavy Water Metabolic Labeling

Journal of Proteome Research ◽

10.1021/acs.jproteome.0c00873 ◽

2021 ◽

Author(s):

Rovshan G. Sadygov

Keyword(s):

Heavy Water ◽

Protein Turnover ◽

Metabolic Labeling

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Identification of an N-acetylneuraminic acid-presenting bacteria isolated from a human microbiome

Scientific Reports ◽

10.1038/s41598-021-83875-w ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Zhen Han ◽

Peter S. Thuy-Boun ◽

Wayne Pfeiffer ◽

Vincent F. Vartabedian ◽

Ali Torkamani ◽

...

Keyword(s):

Pathogenic Bacteria ◽

Human Microbiome ◽

Host Immunity ◽

Rrna Gene ◽

Metabolic Labeling ◽

Selective Labeling ◽

Fecal Microbiome ◽

Acetylneuraminic Acid ◽

Intestinal Mucus ◽

Microbial Systems

AbstractN-Acetylneuraminic acid is the most abundant sialic acid (SA) in humans and is expressed as the terminal sugar on intestinal mucus glycans. Several pathogenic bacteria harvest and display host SA on their own surfaces to evade Siglec-mediated host immunity. While previous studies have identified bacterial enzymes associated with SA catabolism, no reported methods permit the selective labeling, tracking, and quantitation of SA-presenting microbes within complex multi-microbial systems. We combined metabolic labeling, click chemistry, 16S rRNA gene, and whole-genome sequencing to track and identify SA-presenting microbes from a cultured human fecal microbiome. We isolated a new strain of Escherichia coli that incorporates SA onto its own surface and encodes for the nanT, neuA, and neuS genes necessary for harvesting and presenting SA. Our method is applicable to the identification of SA-presenting bacteria from human, animal, and environmental microbiomes, as well as providing an entry point for the investigation of surface-expressed SA-associated structures.

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Quantification of histone modifications using 15N metabolic labeling

Methods ◽

10.1016/j.ymeth.2013.02.004 ◽

2013 ◽

Vol 61 (3) ◽

pp. 236-243 ◽

Cited By ~ 10

Author(s):

Chunchao Zhang ◽

Yifan Liu ◽

Philip C. Andrews

Keyword(s):

Histone Modifications ◽

Metabolic Labeling

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Interrogating Plant-Microbe Interactions with Chemical Tools: Click Chemistry Reagents for Metabolic Labeling and Activity-Based Probes

Molecules ◽

10.3390/molecules26010243 ◽

2021 ◽

Vol 26 (1) ◽

pp. 243

Author(s):

Vivian S. Lin

Keyword(s):

Plant Growth ◽

Immune Responses ◽

Click Chemistry ◽

Chemical Biology ◽

Molecular Mechanisms ◽

Metabolic Labeling ◽

Beneficial Microbes ◽

Plant Hosts ◽

Microbe Interactions ◽

Growth Metabolism

Continued expansion of the chemical biology toolbox presents many new and diverse opportunities to interrogate the fundamental molecular mechanisms driving complex plant–microbe interactions. This review will examine metabolic labeling with click chemistry reagents and activity-based probes for investigating the impacts of plant-associated microbes on plant growth, metabolism, and immune responses. While the majority of the studies reviewed here used chemical biology approaches to examine the effects of pathogens on plants, chemical biology will also be invaluable in future efforts to investigate mutualistic associations between beneficial microbes and their plant hosts.

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