Beyond Mass Spectrometry – Key Proteomics Vocabulary

Vocabulary flashcards covering key terms and concepts from the lecture on emerging proteomics techniques beyond mass spectrometry.

Proteomics

The large-scale study and characterization of all proteins (the proteome) produced by a cell, tissue, or organism.

Mass Spectrometry (MS)

An analytical technique that ionizes chemical species and separates the ions based on their mass-to-charge ratio to identify and characterize molecules such as peptides and proteins.

Bottom-up Mass Spectrometry (BU-MS)

A proteomics workflow in which proteins are enzymatically digested into short peptides that are then analyzed by MS to infer the original protein sequences.

Top-down Mass Spectrometry (TD-MS)

An MS approach that analyzes intact proteins without prior digestion, allowing direct detection of sequence variants and post-translational modifications, typically limited to proteins <70 kDa.

Edman Degradation

A classical chemical method for sequencing peptides by sequentially removing and identifying N-terminal amino acids; accurate but slow and limited to ~30 residues.

Post-translational Modification (PTM)

Chemical alteration of a protein after translation—such as phosphorylation, glycosylation, methylation—that diversifies protein structure and function.

Isoform

A distinct version of a protein produced from the same gene via alternative splicing, proteolytic cleavage, somatic recombination, or PTMs.

Dynamic Range (in MS)

The span between the highest and lowest detectable peptide/protein abundances; typically ~5 orders of magnitude for Orbitrap instruments, whereas biological samples can span 12.

Sensitivity (in proteomics)

The minimum quantity of protein required for reliable detection; traditional MS often needs attomole-to-femtomole (millions-to-billions of molecules) amounts.

Central Dogma

The flow of genetic information from DNA to RNA (transcription) and from RNA to protein (translation).

Open Reading Frame (ORF)

A continuous stretch of codons in mRNA that begins with a start codon and ends at a stop codon, potentially encoding a protein.

Alternative Splicing

A regulatory process that joins different combinations of exons to produce multiple mRNA—and hence protein—isoforms from one gene.

Long-read Sequencing

DNA or RNA sequencing technologies (e.g., PacBio, Oxford Nanopore) that generate reads of thousands of bases, aiding full-length isoform identification.

CITE-seq

Cellular Indexing of Transcriptomes and Epitopes by Sequencing; a method that uses DNA-barcoded antibodies and droplet sequencing to measure RNA and surface proteins simultaneously in single cells.

Fluorescent Protein Fingerprinting

A single-molecule strategy that labels specific amino acids with fluorophores and records fluorescence changes (e.g., during Edman cycles or ClpXP translocation) to partially sequence peptides.

Single-molecule FRET

Fluorescence Resonance Energy Transfer measured at the single-molecule level to monitor distances between donor and acceptor dyes, used here to read peptide fingerprints.

Nanopore

A nanometer-scale hole in a membrane through which ionic current passes; analyte translocation modulates the current, enabling single-molecule detection.

Subnanopore

An ultrasmall (<1 nm waist) solid-state nanopore that can resolve features roughly the size of individual amino acids in unfolded proteins.

5D Fingerprinting

Nanopore analysis that extracts five properties—shape, volume, charge, rotational diffusion coefficient, and dipole moment—of single folded proteins from current modulations.

Diffusion Capacitance

The effective ‘reservoir’ of molecules near a sensor that governs capture rate; minimizing it (e.g., by positioning cells close to nanopores) enhances detection sensitivity.

Dielectric Noise

Current noise originating from the capacitance of the nanopore membrane and measurement circuitry, dominant above ~1 kHz and limiting high-bandwidth recordings.

Random-Forest Model (in nanopore reads)

A machine-learning approach using multiple decision trees to predict amino-acid identities from complex nanopore blockade signals.

ClpXP

An AAA+ protease complex that unfolds and translocates proteins, enabling controlled peptide passage through nanopores or FRET zones for fingerprinting.

AAA+ Protease

A family of ATP-dependent enzymes that remodel or degrade proteins by unfolding and threading them through a proteolytic chamber.

Peptide Mass Fingerprinting

Identifying a protein by matching experimentally measured peptide masses (from protease digestion) to theoretical masses in a database.

Attomole (amol)

10⁻¹⁸ moles; roughly 600,000 molecules of a 50-kDa protein.

Femtomole (fmol)

10⁻¹⁵ moles; about 600 million molecules of a 50-kDa protein.

Electric Field Focusing (in nanopores)

Confinement of the electric field to a small region (≈1 nm) near a pore waist, enhancing spatial resolution of current measurements.

Hydrodynamic Diameter

Effective diameter of a molecule moving in solution; when comparable to pore size, translocation speed decreases dramatically.

SDS Denaturation

Use of sodium dodecyl sulfate to linearize proteins and coat them with negative charge, facilitating uniform electrophoretic translocation through nanopores.