Analysis of Peptides by Denaturing Ultrafiltration and LC-MALDI-TOF-MS

An, Yanming; Goldman, Radoslav

doi:10.1007/978-1-4614-7209-4_2

Cited by 6 publications

(3 citation statements)

References 22 publications

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“…The advantages of different methods have previously been compared for peptide recovery . The most used peptide enrichment method is centrifugal ultrafiltration, which uses molecular weight cutoff (MWCO) membranes to separate low and high molecular weight peptides . This method is fast, easy to perform and can identify many unique peptides in serum and plasma. , However, peptides with molecular weights close to the cutoff point may be lost, such as apolipoproteins, and some “sticky” lipophilic peptides might be lost by binding to the membranes.…”

Section: Methodologies For Peptidomics Analysismentioning

confidence: 99%

Peptidomics: A Review of Clinical Applications and Methodologies

et al. 2021

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Improvements in both liquid chromatography (LC) and mass spectrometry (MS) instrumentation have greatly enhanced proteomic and small molecule metabolomic analysis in recent years. Less focus has been on the improved capability to detect and quantify small bioactive peptides, even though the exact sequences of the peptide species produced can have important biological consequences. Endogenous bioactive peptide hormones, for example, are generated by the targeted and regulated cleavage of peptides from their prohormone sequence. This process may include organ specific variants, as proglucagon is converted to glucagon in the pancreas but glucagon-like peptide-1 (GLP-1) in the small intestine, with glucagon raising, whereas GLP-1, as an incretin, lowering blood glucose. Therefore, peptidomics workflows must preserve the structure of the processed peptide products to prevent the misidentification of ambiguous peptide species. The poor in vivo and in vitro stability of peptides in biological matrices is a major factor that needs to be considered when developing methods to study them. The bioinformatic analysis of peptidomics data sets requires the inclusion of specific posttranslational modifications, which are critical for the function of many bioactive peptides. This review aims to discuss and contrast the various extraction, analytical, and bioinformatics approaches used for human peptidomics studies in a multitude of matrices.

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Section: Methodologies For Peptidomics Analysismentioning

confidence: 99%

Peptidomics: A Review of Clinical Applications and Methodologies

et al. 2021

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“…As an alternative approach, the fact that the majority of the high abundance proteins in serum are of comparatively high molecular weight has been used as a characteristic upon which to base a depletion protocol. Ultrafi ltration of serum with fi lters with molecular weight cut-offs of 50 and 30 kDa has been applied as a methodology in order to allow for the study of low abundance proteins/peptides [ 3 ]. Such protocols commonly dilute the serum with acidic solutions (2 % trifl uoroacetic acid for example) in order to dissociate peptides and proteins bound to albumin.…”

Section: Sample Preparation For Protein/peptides For Protein Profi Lingmentioning

confidence: 99%

MALDI Profiling and Applications in Medicine

Dudley

2014

Advances in Experimental Medicine and Biology

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Matrix-assisted laser desorption ionisation (MALDI) mass spectrometry allows for the rapid profiling of different biomolecular species from both biofluids and tissues. Whilst originally focussed upon the analysis of intact proteins and peptides, MALDI mass spectrometry has found further applications in lipidomic analysis, genotyping, micro-organism identification, biomarker discovery and metabolomics. The combining of multiple profiles data from differing locations across a sample, furthermore, allows for spatial distribution of biomolecules to be established utilising imaging MALDI analysis. This chapter discusses the MALDI process, its usual applications in the field of protein identification via peptide mass fingerprinting before focusing upon advances in the application of the profiling potential of MALDI mass spectrometry and its various applications in biomedicine.

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“…Another study reported a method that coupled denaturing ultrafiltration with reverse phase fractionation and MS for characterization of low‐molecular‐weight proteins and peptides in serum and plasma samples . The enriched peptides were analyzed by RP‐LC combined with MALDI‐MS/MS characterization of the analytes, resulting in identification of 250 native peptides from 50 different proteins .…”

Section: Biomarker Discoverymentioning

confidence: 99%

Coupling enrichment methods with proteomics for understanding and treating disease

Kumar

Baycin-Hizal²,

Shiloach

et al. 2015

Proteomics Clinical Apps

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Owing to recent advances in proteomics analytical methods and bioinformatics capabilities there is a growing trend toward using these capabilities for the development of drugs to treat human disease, including target and drug evaluation, understanding mechanisms of drug action, and biomarker discovery. Currently, the genetic sequences of many major organisms are available, which have helped greatly in characterizing proteomes in model animal systems and humans. Through proteomics, global profiles of different disease states can be characterized (e.g. changes in types and relative levels as well as changes in PTMs such as glycosylation or phosphorylation). Although intracellular proteomics can provide a broad overview of physiology of cells and tissues, it has been difficult to quantify the low abundance proteins which can be important for understanding the diseased states and treatment progression. For this reason, there is increasing interest in coupling comparative proteomics methods with subcellular fractionation and enrichment techniques for membranes, nucleus, phosphoproteome, glycoproteome as well as low abundance serum proteins. In this review, we will provide examples of where the utilization of different proteomics-coupled enrichment techniques has aided target and biomarker discovery, understanding the drug targeting mechanism, and mAb discovery. Taken together, these improvements will help to provide a better understanding of the pathophysiology of various diseases including cancer, autoimmunity, inflammation, cardiovascular disease, and neurological conditions, and in the design and development of better medicines for treating these afflictions.

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Analysis of Peptides by Denaturing Ultrafiltration and LC-MALDI-TOF-MS

Cited by 6 publications

References 22 publications

Peptidomics: A Review of Clinical Applications and Methodologies

Peptidomics: A Review of Clinical Applications and Methodologies

MALDI Profiling and Applications in Medicine

Coupling enrichment methods with proteomics for understanding and treating disease

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