Research method
Mass Spectrometry Proteomics
7 studies
Mass-spectrometry proteomics identifies and, when labelled or quantified, counts peptides from a protein mixture after proteolytic digestion. Outputs range from a subcellular inventory (who is on this membrane face) to site maps of modifications (ubiquitin, lysine crotonylation) to subunit stoichiometry of a purified complex. An identified peptide is not a phenotype, and genome papers in this list are not proteomic experiments.
Cell biologists use MS when fractionation or antibodies cannot resolve a compartment or a modification site. It answers 'which proteins or modified lysines are in this sample?' The main limitation is that enrichment is not functional proof, and several lexicon hits here are genomes or transcriptomes that never ran a mass spectrometer on peptides.
Studies
7
Findings
5
7 supporting · 0 challenging · 0 qualifying citations
Open tensions
2
Currently
What we know
- APEX2 proximity labelling in living cells defined cytosol-facing proteomes of the outer mitochondrial membrane and the ER membrane: expected functional classes plus many additions (22 OMM proteins; 72 ERM proteins), beyond classical fractionation. The inventory is not a full functional validation of every newly enriched protein.
- Purified caveolar coats analysed by quantitative composition showed caveolins and cavins as one complex excluding EHD2/pacsin 2, with Cavin 1 a core component and evidence for trimers. MS-backed stoichiometry is a biochemical model, not a dynamics movie in every cell type.
- Rab5 monoubiquitination sites were mapped at K116, K140 and K165; chemically monoubiquitinated Rab5 then showed that K140/K165 downregulate the endocytic pathway via disrupted effectors and nucleotide conversion, while K116 has little localisation effect. Site-resolved MS (or MS-informed chemistry) here is a modification map, not an identification of the endogenous E3 ligase.
- In rice, MS plus ChIP-seq mapped histone lysine butyrylation and crotonylation: many Kcr sites on H3/H4, with Kbu/Kcr resembling acetylation distribution and changing with metabolic stress. Occupancy dynamics are not proof that every site is required for a named stress phenotype.
- Three papers are false-positive lexicon hits for proteomics. Naegleria fowleri work assembled an ~30 Mb AT-rich genome and listed candidate pathogenicity genes; Aspergillus comparative genomics reported ~29–36 Mb genomes with 9,113–13,553 genes (~20% Aspergillaceae-specific; section Nigri ~1,800 unique genes); Plasmodium falciparum IDC transcriptome was hourly expression across a ~48-hour blood-stage cycle. Those are DNA or RNA catalogues, not peptide spectra.
Largest unresolved question
Genuine MS designs still disagree about what is being counted. APEX2 inventories membrane-face neighbours in living cells (22 OMM; 72 ERM); caveolar work quantifies a purified coat; Rab5 work maps three ubiquitin lysines; rice work maps histone acylations under stress. A 'proteomics paper' can be spatial, stoichiometric, or epigenetic, and those outputs are not interchangeable.
Procedure
How it works
The main stages of the method. Specific protocols and analysis choices vary between studies.
Digest the protein mixture
Proteins from a lysate, a proximity-labelled compartment, or a purified complex are cut into peptides. What never entered the digest cannot be identified later.
Separate and ionise peptides
Peptides are usually chromatographed then ionised into a mass spectrometer. Genome or transcriptome papers tagged for this method never took this step.
Match spectra to sequences
Fragment ions are identified as peptides. An identified peptide is a sequence in this sample, not a phenotype and not functional proof that the protein acts on that membrane face.
Quantify proteins, sites, or stoichiometry
Outputs range from a subcellular inventory to ubiquitin or crotonylation site maps to subunit ratios of a coat. Site-specific chemistry, not a bulk spectral count, is what distinguishes a Rab5 lysine that downregulates endocytosis from one that does not.
Treat an identified peptide as inventory, not a phenotype
MS proteomics answers which proteins or modified lysines are in this sample. Enrichment is a location hypothesis; candidate gene lists from sequenced genomes are not peptide identifications.
Common misconceptions
If APEX2 enriches a protein on the OMM or ERM, that protein's function on that face is known.
The maps capture expected classes and add 22 OMM and 72 ERM proteins beyond classical fractionation. Enrichment is a location hypothesis, not functional validation of each new protein.
Detecting ubiquitin on Rab5 is enough to say the GTPase is off.
K140 and K165 modifications downregulate endocytosis; K116 has little localisation effect. Site-specific chemistry, not a bulk ubiquitin spectral count, made that distinction.
A pathogenicity-factor 'proteome' of Naegleria was measured because the paper is tagged for mass spectrometry.
The study sequenced and assembled an ~30 Mb AT-rich genome and nominated candidate genes. Those candidates still need functional validation in infection models and are not peptide identifications.
Related
Claim ledger
What the evidence shows
Drawn from 7 studies in this library. Mix labels say which citation roles are present; they are not a strength score. Supports means evidence for a finding; Challenges means evidence against a stated position; Qualifies marks scope.
APEX2 proximity labelling in living cells defined cytosol-facing proteomes of the outer mitochondrial membrane and the ER membrane: expected functional classes plus many additions (22 OMM proteins; 72 ERM proteins), beyond classical fractionation. The inventory is not a full functional validation of every newly enriched protein.
Purified caveolar coats analysed by quantitative composition showed caveolins and cavins as one complex excluding EHD2/pacsin 2, with Cavin 1 a core component and evidence for trimers. MS-backed stoichiometry is a biochemical model, not a dynamics movie in every cell type.
Rab5 monoubiquitination sites were mapped at K116, K140 and K165; chemically monoubiquitinated Rab5 then showed that K140/K165 downregulate the endocytic pathway via disrupted effectors and nucleotide conversion, while K116 has little localisation effect. Site-resolved MS (or MS-informed chemistry) here is a modification map, not an identification of the endogenous E3 ligase.
In rice, MS plus ChIP-seq mapped histone lysine butyrylation and crotonylation: many Kcr sites on H3/H4, with Kbu/Kcr resembling acetylation distribution and changing with metabolic stress. Occupancy dynamics are not proof that every site is required for a named stress phenotype.
Three papers are false-positive lexicon hits for proteomics. Naegleria fowleri work assembled an ~30 Mb AT-rich genome and listed candidate pathogenicity genes; Aspergillus comparative genomics reported ~29–36 Mb genomes with 9,113–13,553 genes (~20% Aspergillaceae-specific; section Nigri ~1,800 unique genes); Plasmodium falciparum IDC transcriptome was hourly expression across a ~48-hour blood-stage cycle. Those are DNA or RNA catalogues, not peptide spectra.
- Pathogenicity factors in Naegleria fowleri
- Aspergillus genomic diversity
- How does malaria’s blood-stage transcriptome unfold?
Study Role Design N Population Outcome Pathogenicity factors in Naegleria fowleri Supports Computational / modellingDe novo genome sequencing plus proteomics to nominate N. fowleri pathogenicity factors ~29.6 Mb assembled nuclear genome (diploid estimate ~66 Mb); >500 million reads — genome resource study Naegleria fowleri (compared with non-pathogenic N. gruberi) Candidate pathogenicity gene sets from genome/proteome comparison Aspergillus genomic diversity Supports Computational / modellingComparative genomics of Aspergillus genomes in MycoCosm Multi-genome Aspergillus comparison (≈29–36 Mb; 9,113–13,553 genes) — genome count not a single primary N in stored summary Aspergillus / Aspergillaceae genomes Genome size, gene content, and clade-specific gene expansions How does malaria’s blood-stage transcriptome unfold? Supports Animal / in-vitroHourly IDC transcriptome of P. falciparum across the ~48-hour erythrocytic cycle Time-series parasite culture transcriptome — no single sample N Plasmodium falciparum during the intraerythrocytic developmental cycle Stage-ordered periodic gene expression across the IDC
Debates
Tensions and limits
Some items are genuine disagreements on the same question. Others mark different assays, populations, or outcomes.
Genuine MS designs still disagree about what is being counted. APEX2 inventories membrane-face neighbours in living cells (22 OMM; 72 ERM); caveolar work quantifies a purified coat; Rab5 work maps three ubiquitin lysines; rice work maps histone acylations under stress. A 'proteomics paper' can be spatial, stoichiometric, or epigenetic, and those outputs are not interchangeable.
Genuine MS designs still disagree about what is being counted. APEX2 inventories membrane-face neighbours in living cells (22 OMM; 72 ERM); caveolar work quantifies a purified coat; Rab5 work maps three ubiquitin lysines; rice work maps histone acylations under stress. A 'proteomics paper' can be spatial, stoichiometric, or epigenetic, and those outputs are not interchangeable.
- Mapping the faces of mitochondria and ER
- What proteins build the caveolar coat?
- How does monoubiquitin turn down Rab5?
- Histone acylations respond to plant stress
Study Role Design N Population Outcome Mapping the faces of mitochondria and ER Supports Animal / in-vitroAPEX2 proximity labeling of cytosol-facing OMM and ER membranes in living HEK 293T cells SILAC proteomic maps with two replicates per membrane — cell-resource study HEK 293T cells expressing OMM/ERM APEX2 fusions Proteomes of cytosol-facing outer mitochondrial and ER membranes What proteins build the caveolar coat? Supports Animal / in-vitroBiochemical purification and ultrastructure of caveolar coat proteins Structural/biochemical coat stoichiometry study — no single sample N Caveolar coat complexes (caveolins/cavins) Molecular composition and ultrastructure of the caveolar coat How does monoubiquitin turn down Rab5? Supports Animal / in-vitroMapped Rab5 monoubiquitination sites and tested chemically monoubiquitinated Rab5 function Cell-biochemistry of Rab5 ubiquitin sites K116/K140/K165 — no single sample N Cultured cells expressing Rab5 variants Site-specific monoubiquitination downregulating Rab5 endocytic signaling Histone acylations respond to plant stress Supports Animal / in-vitroMS and ChIP-seq mapping of histone Kbu/Kcr in rice vs H3K9ac under metabolic stress Plant chromatin proteomics/ChIP-seq study — no single primary analytic N in stored text Rice (and related plants) under stress conditions Distribution and stress dynamics of histone butyrylation/crotonylation
PaperFren reads this as a limit on how far one study travels — different assays, populations, or outcomes — not a forced fight between papers.
Naegleria and Aspergillus genomes and the Plasmodium IDC transcriptome should not be cited as mass-spectrometry evidence at all. Candidate pathogenicity gene lists and periodic RNA profiles are sequence resources that still need protein-level or infection-model tests.
Naegleria and Aspergillus genomes and the Plasmodium IDC transcriptome should not be cited as mass-spectrometry evidence at all. Candidate pathogenicity gene lists and periodic RNA profiles are sequence resources that still need protein-level or infection-model tests.
- Pathogenicity factors in Naegleria fowleri
- Aspergillus genomic diversity
- How does malaria’s blood-stage transcriptome unfold?
Study Role Design N Population Outcome Pathogenicity factors in Naegleria fowleri Supports Computational / modellingDe novo genome sequencing plus proteomics to nominate N. fowleri pathogenicity factors ~29.6 Mb assembled nuclear genome (diploid estimate ~66 Mb); >500 million reads — genome resource study Naegleria fowleri (compared with non-pathogenic N. gruberi) Candidate pathogenicity gene sets from genome/proteome comparison Aspergillus genomic diversity Supports Computational / modellingComparative genomics of Aspergillus genomes in MycoCosm Multi-genome Aspergillus comparison (≈29–36 Mb; 9,113–13,553 genes) — genome count not a single primary N in stored summary Aspergillus / Aspergillaceae genomes Genome size, gene content, and clade-specific gene expansions How does malaria’s blood-stage transcriptome unfold? Supports Animal / in-vitroHourly IDC transcriptome of P. falciparum across the ~48-hour erythrocytic cycle Time-series parasite culture transcriptome — no single sample N Plasmodium falciparum during the intraerythrocytic developmental cycle Stage-ordered periodic gene expression across the IDC
PaperFren reads this as a limit on how far one study travels — different assays, populations, or outcomes — not a forced fight between papers.
Timeline
How understanding moved
Study years are when the paper was published. Evidence edits are dated changes to this page's claims. Explanations are when PaperFren added a Discovery — not a claim that the science happened that day.
2026
Concept page published
Mass Spectrometry Proteomics
Change log
What changed
Dated edits to this page's evidence: studies added or removed from a claim, claims added or withdrawn, and new explanations tagged here. Coverage edits are distinct from reviewed research changes and corrections. Dates record PaperFren updates; study publication dates appear in the timeline.
- Coverage updated · Concept page published
Papers
7 studies in this library bear on Mass Spectrometry Proteomics, ordered by citations.
- How does malaria’s blood-stage transcriptome unfold?
Plasmodium falciparum’s 48-hour intraerythrocytic cycle was profiled hourly, revealing tightly staged gene-expression programs after RBC invasion.
- Aspergillus genomic diversity
Comparative genomics shows Aspergillus species share genome size but harbor large lineage-specific gene sets.
- Mapping the faces of mitochondria and ER
APEX2 proximity biotinylation yields high-quality proteomic maps of cytosol-facing outer mitochondrial and ER membranes in living human cells.
- What proteins build the caveolar coat?
Caveolins and cavins purify as a caveolar coat complex with Cavin 1 as a core trimeric component.
- Histone acylations respond to plant stress
Rice histone butyrylation and crotonylation mark active chromatin and shift under starvation/submergence.
- Pathogenicity factors in Naegleria fowleri
Whole-genome analysis of N. fowleri highlights candidate pathogenicity factors for brain infection.
- How does monoubiquitin turn down Rab5?
Rab5 is monoubiquitinated at specific lysines; ubiquitin at K140/K165 disrupts effector binding and downregulates endocytic trafficking.
Compare studies
Select 2–10 studies. Design and N are labels, not a ranking.
Nothing selected yet.
Questions
What is still open
Genuine MS designs still disagree about what is being counted. APEX2 inventories membrane-face neighbours in living cells (22 OMM; 72 ERM); caveolar work quantifies a purified coat; Rab5 work maps three ubiquitin lysines; rice work maps histone acylations under stress. A 'proteomics paper' can be spatial, stoichiometric, or epigenetic, and those outputs are not interchangeable.
Naegleria and Aspergillus genomes and the Plasmodium IDC transcriptome should not be cited as mass-spectrometry evidence at all. Candidate pathogenicity gene lists and periodic RNA profiles are sequence resources that still need protein-level or infection-model tests.
Ask PaperFren about Mass Spectrometry Proteomics
Study this conceptflashcards and short-answer questions
Why can APEX2 list proteins that classical mitochondrial fractionation misses, and what can it still not claim?
Proximity biotinylation in living cells labels cytosol-facing OMM and ERM neighbours (22 and 72 proteins beyond expected classes). It does not prove each enriched protein's function or rule out labelling of transient visitors.
Caveolin–cavin coats exclude EHD2/pacsin 2. How is that a different MS question from mapping rice H3/H4 crotonylation sites?
Coat work is subunit stoichiometry of a purified complex (Cavin 1 core, trimers). Rice work is a modification atlas that resembles acetylation and changes with metabolic stress. One counts protein copies in a particle; the other counts modified lysines on histones.
Chemically monoubiquitinated Rab5 at K140/K165 versus K116 gave different endocytic outcomes. Why was mapping sites necessary before that experiment?
Without site maps (K116, K140, K165), a researcher would not know which lysines to modify. The chemistry tests those mapped sites; it still does not identify the endogenous E3 ligase.
Plasmodium IDC expression is highly periodic over ~48 hours. Why is that a poor example of mass-spectrometry proteomics?
The dataset is an hourly transcriptome from merozoite invasion through schizont stages, not a peptide inventory. Periodicity of mRNA is not a protein-level MS result and does not itself deliver a drug.