Metabolomic studies allow a deeper understanding of the processes of a given ecological community than nucleic acid–based surveys alone. In the case of the gut microbiota, a metabolic profile of, for example, a fecal sample provides details about the function and interactions within the distal region of the gastrointestinal tract, and such a profile can be generated in a number of different ways. This unit elaborates on the use of 1D1H NMR spectroscopy as a commonly used method to characterize small‐molecule metabolites of the fecal metabonome (meta‐metabolome). We describe a set of protocols for the preparation of fecal water extraction, storage, scanning, measurement of pH, and spectral processing and analysis. We also compare the effects of various sample storage conditions for processed and unprocessed samples to provide a framework for comprehensive analysis of small molecules from stool‐derived samples. © 2020 Wiley Periodicals LLC
Glycosylated proteins, namely glycoproteins and proteoglycans (collectively called glycoconjugates), are indispensable in a variety of biological processes. The functions of many glycoconjugates are regulated by their interactions with another group of proteins known as lectins. In order to understand the biological functions of lectins and their glycosylated binding partners, one must obtain these proteins in pure form. The conventional protein purification methods often require long times, elaborate infrastructure, costly reagents, and large sample volumes. To minimize some of these problems, we recently developed and validated a new method termed capture and release (CaRe). This method is time‐saving, precise, inexpensive, and it needs a relatively small sample volume. In this approach, targets (lectins and glycoproteins) are captured in solution by multivalent ligands called target capturing agents (TCAs). The captured targets are then released and separated from their TCAs to obtain purified targets. Application of the CaRe method could play an important role in discovering new lectins and glycoconjugates. © 2020 Wiley Periodicals LLC.
- PAR ID:
- 10238502
- Publisher / Repository:
- Wiley Blackwell (John Wiley & Sons)
- Date Published:
- Journal Name:
- Current Protocols in Protein Science
- Volume:
- 101
- Issue:
- 1
- ISSN:
- 1934-3655
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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Basic Protocol 1 : Cell transfection (optional)Support Protocol : Preparation of degassed lysis buffersBasic Protocol 2 : Cellular extraction in anaerobic conditionsBasic Protocol 3 : Enrichment and activity assay of specific PTPsAlternate Protocol : Measurement of active PTPs via direct cysteinyl labeling -
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This article was corrected on 18 July 2022. See the end of the full text for details.
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