Assessment of Label-Free Quantification in Discovery Proteomics and Impact of Technological Factors and Natural Variability of Protein Abundance

Shweiki, Mhd Rami Al; Mönchgesang, Susann; Majovsky, Petra; Thieme, Domenika; Trutschel, Diana; Hoehenwarter, Wolfgang

doi:10.1021/acs.jproteome.6b00645

Cited by 95 publications

(74 citation statements)

References 44 publications

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“…As previously described [ 58 ], we used MaxQuant iBAQ [ 35 , 72 ] to discern minor and major proteins. The peptide yield of the previously localized extra-crystalline matrix protein lustrin A [ 12 , 13 ], predominantly identified in the acid-insoluble fractions of nacre, was not particularly affected when the shell was treated with sodium hypochlorite (Additional file 4 : Table S2), indicating that our relatively mild washing most probably not did destroy the extra-crystalline matrix to an appreciable extent.…”

Section: Resultsmentioning

confidence: 99%

In-depth proteomic analyses of Haliotis laevigata (greenlip abalone) nacre and prismatic organic shell matrix

et al. 2018

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BackgroundThe shells of various Haliotis species have served as models of invertebrate biomineralization and physical shell properties for more than 20 years. A focus of this research has been the nacreous inner layer of the shell with its conspicuous arrangement of aragonite platelets, resembling in cross-section a brick-and-mortar wall. In comparison, the outer, less stable, calcitic prismatic layer has received much less attention. One of the first molluscan shell proteins to be characterized at the molecular level was Lustrin A, a component of the nacreous organic matrix of Haliotis rufescens. This was soon followed by the C-type lectin perlucin and the growth factor-binding perlustrin, both isolated from H. laevigata nacre, and the crystal growth-modulating AP7 and AP24, isolated from H. rufescens nacre. Mass spectrometry-based proteomics was subsequently applied to to Haliotis biomineralization research with the analysis of the H. asinina shell matrix and yielded 14 different shell-associated proteins. That study was the most comprehensive for a Haliotis species to date.MethodsThe shell proteomes of nacre and prismatic layer of the marine gastropod Haliotis laevigata were analyzed combining mass spectrometry-based proteomics and next generation sequencing.ResultsWe identified 297 proteins from the nacreous shell layer and 350 proteins from the prismatic shell layer from the green lip abalone H. laevigata. Considering the overlap between the two sets we identified a total of 448 proteins. Fifty-one nacre proteins and 43 prismatic layer proteins were defined as major proteins based on their abundance at more than 0.2% of the total. The remaining proteins occurred at low abundance and may not play any significant role in shell fabrication. The overlap of major proteins between the two shell layers was 17, amounting to a total of 77 major proteins.ConclusionsThe H. laevigata shell proteome shares moderate sequence similarity at the protein level with other gastropod, bivalve and more distantly related invertebrate biomineralising proteomes. Features conserved in H. laevigata and other molluscan shell proteomes include short repetitive sequences of low complexity predicted to lack intrinsic three-dimensional structure, and domains such as tyrosinase, chitin-binding, and carbonic anhydrase. This catalogue of H. laevigata shell proteins represents the most comprehensive for a haliotid and should support future efforts to elucidate the molecular mechanisms of shell assembly.Electronic supplementary materialThe online version of this article (10.1186/s12953-018-0139-3) contains supplementary material, which is available to authorized users.

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Section: Resultsmentioning

confidence: 99%

In-depth proteomic analyses of Haliotis laevigata (greenlip abalone) nacre and prismatic organic shell matrix

et al. 2018

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“…The customized bioproduction would take several months, be expensive, and success would be uncertain. Therefore, a label‐free approach was developed, which has been already shown to be promising 41 …”

Section: Resultsmentioning

confidence: 99%

Human prorenin determination by hybrid immunocapture liquid chromatography/mass spectrometry: A mixed‐solvent‐triggered digestion utilizing D‐optimal design

Burdman

Burckhardt

2020

Rapid Comm Mass Spectrometry

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Rationale Human prorenin, representing the precursor of mature renin, has been discussed as a potential biomarker, e.g. in diagnosing primary hyperaldosteronism or diabetes‐induced nephropathy. Currently, only immunoassays are available for prorenin quantification. As the similarity of prorenin to active renin impedes its accurate determination by immunoassay, mass spectrometry appears as an accurate alternative for differentiation of that protein. Methods Immunoaffinity purification plus a mixed‐solvent‐triggered digestion was combined with liquid chromatography/mass spectrometry (LC/MS) to enable a fast, sensitive, and less laboratory‐intensive approach to the quantification of prorenin. Statistical experimental planning, which is known as Design of Experiments (DOE), was used to identify the optimal conditions for the generation of the signature peptides within a manageable number of experiments. The efficiency of the mixed‐solvent‐triggered digestion by trypsin was investigated using four different organic solvents: acetonitrile, acetone, tetrahydrofuran and methanol. Results By utilizing a D‐optimal design, we found that the optimal mixed‐solvent type for the generation of both signature peptides was acetonitrile at a concentration of 84% and an incubation temperature of 16°C. Using the mixed‐solvent‐triggered digestion, the procedure time allowed a fast analysis of active renin and prorenin with a short digestion time of 98 min. This optimized mixed‐solvent‐triggered digestion procedure was applied to detect renin and prorenin successfully in human plasma by the newly developed hybrid approach. Conclusions The identification of unique surrogates for human prorenin enabled the mass spectrometric differentiation between the two similar proteins. The novel hybrid approach successfully proved its ability to purify, detect and distinguish between prorenin and active renin in human plasma.

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“…The MaxQuant 26 software suite is used to analyze the MS data in this protocol, but commercial applications 35 are also available for phosphopeptide identification and quantification. For phosphopeptide identification, a localization probability cutoff is applied.…”

Section: Discussionmentioning

confidence: 99%

Phosphopeptide Enrichment Coupled with Label-free Quantitative Mass Spectrometry to Investigate the Phosphoproteome in Prostate Cancer

Cheng

Graeber

et al. 2018

JoVE

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Phosphoproteomics involves the large-scale study of phosphorylated proteins. Protein phosphorylation is a critical step in many signal transduction pathways and is tightly regulated by kinases and phosphatases. Therefore, characterizing the phosphoproteome may provide insights into identifying novel targets and biomarkers for oncologic therapy. Mass spectrometry provides a way to globally detect and quantify thousands of unique phosphorylation events. However, phosphopeptides are much less abundant than non-phosphopeptides, making biochemical analysis more challenging. To overcome this limitation, methods to enrich phosphopeptides prior to the mass spectrometry analysis are required. We describe a procedure to extract and digest proteins from tissue to yield peptides, followed by an enrichment for phosphotyrosine (pY) and phosphoserine/threonine (pST) peptides using an antibody-based and/or titanium dioxide (TiO2)-based enrichment method. After the sample preparation and mass spectrometry, we subsequently identify and quantify phosphopeptides using liquid chromatography-mass spectrometry and analysis software.

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Assessment of Label-Free Quantification in Discovery Proteomics and Impact of Technological Factors and Natural Variability of Protein Abundance

Cited by 95 publications

References 44 publications

In-depth proteomic analyses of Haliotis laevigata (greenlip abalone) nacre and prismatic organic shell matrix

In-depth proteomic analyses of Haliotis laevigata (greenlip abalone) nacre and prismatic organic shell matrix

Human prorenin determination by hybrid immunocapture liquid chromatography/mass spectrometry: A mixed‐solvent‐triggered digestion utilizing D‐optimal design

Phosphopeptide Enrichment Coupled with Label-free Quantitative Mass Spectrometry to Investigate the Phosphoproteome in Prostate Cancer

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