15  Proteomics

15.1 From Transcript to Protein

15.1.1 What Is Proteomics?

Proteomics is the large-scale study of proteins—their structures, functions, and interactions.

The “ome” suffix:

  • Genome = all DNA

  • Transcriptome = all RNA

  • Proteome = all proteins

Think of the proteome as all the workers actually doing jobs in your cells right now!

15.1.2 Why Study Proteins?

Proteins are where the action is:

  • Genes are instructions

  • RNA is the messenger

  • Proteins do the actual work!

Understanding proteins helps us:

  • Know how cells function

  • Understand diseases

  • Develop better medicines

  • Design new therapies

15.2 The Journey: DNA → RNA → Protein

15.2.1 Quick Review

1. Transcription: DNA → RNA

  • Makes mRNA copy of gene

  • In the nucleus

2. Translation: RNA → Protein

  • Ribosomes read mRNA

  • tRNAs bring amino acids

  • Amino acids link together

  • Forms protein chain

3. Protein Folding: Linear chain → 3D structure

  • Protein folds into specific 3D shape

  • Shape determines function

  • “Structure equals function”!

15.2.2 The Proteome Is More Complex Than the Transcriptome

Why?

  • One gene → multiple RNA variants (alternative splicing)

  • One RNA → one protein chain

  • BUT one protein chain → multiple modified forms!

Post-translational modifications (we’ll discuss next) create even MORE diversity!

Result:

  • ~20,000 genes

  • ~100,000 RNA variants

  • Over 1 million different protein forms!

15.3 Post-Translational Modifications (PTMs)

15.3.1 Proteins Get Modified After They’re Made

After a protein is made, it can be modified in many ways!

Post-translational modifications (PTMs) = Chemical changes to proteins after translation

Think of it like:

  • Making a basic clay pot (translation)

  • Then decorating it (PTMs)

  • The decorations change how it’s used!

15.3.2 Common Types of PTMs

15.3.3 1. Phosphorylation

What happens: Adding a phosphate group (PO₄) to amino acids

Where: Usually on serine, threonine, or tyrosine amino acids

Effects:

  • Changes protein shape

  • Can turn protein ON or OFF

  • Very common in cell signaling

Example: Insulin signaling uses lots of phosphorylation to control blood sugar!

15.3.4 2. Methylation

What happens: Adding a methyl group (CH₃) to amino acids

Where: Usually on lysine or arginine

Effects:

  • Controls gene expression (on histones)

  • Affects protein-protein interactions

  • Important for epigenetics!

15.3.5 3. Acetylation

What happens: Adding an acetyl group to amino acids

Where: Usually on lysine

Effects:

  • Loosens chromatin (on histones)

  • Activates some enzymes

  • Opposite of methylation effects (sometimes)

15.3.6 4. Ubiquitination

What happens: Attaching ubiquitin (a small protein) to target protein

Effects:

  • Tags protein for degradation (recycling)

  • Like putting a “trash” label on the protein

  • Controls protein lifespan

15.3.7 5. Glycosylation

What happens: Adding sugar molecules to protein

Where: Usually on asparagine or serine/threonine

Effects:

  • Protects protein from degradation

  • Helps protein folding

  • Important for cell recognition

  • About 50% of human proteins are glycosylated!

15.3.8 6. Proteolytic Cleavage

What happens: Cutting the protein chain into pieces

Effects:

  • Activates some proteins (insulin is made this way!)

  • Inactivates others

  • Removes signaling sequences

  • Irreversible (can’t be undone)

15.3.9 Why PTMs Matter

Creates enormous diversity:

  • One protein can exist in many modified forms

  • Different modifications = different functions

  • Like having one tool that changes based on attachments!

Allows rapid regulation:

  • Adding/removing PTMs is faster than making new proteins

  • Quick response to signals

  • Fine-tuned control

Examples of PTM importance:

  • Cancer: Abnormal phosphorylation drives cancer growth

  • Alzheimer’s: Abnormal protein modifications in brain

  • Diabetes: Insulin signaling requires proper phosphorylation

  • Aging: Accumulated protein modifications over time

15.4 Protein Structure and Function

15.4.1 The Four Levels of Protein Structure

Primary structure (1°):

  • The sequence of amino acids

  • Like letters in a sentence

  • Determined by DNA sequence

Secondary structure (2°):

  • Local folding patterns

  • Alpha helices (spiral staircases)

  • Beta sheets (zigzag ribbons)

  • Held together by hydrogen bonds

Tertiary structure (3°):

  • Overall 3D shape of one protein chain

  • How secondary structures fold together

  • The complete 3D structure

  • Determines function!

Quaternary structure (4°):

  • Multiple protein chains together

  • Not all proteins have this

  • Example: Hemoglobin (4 chains working together)

15.4.2 Structure = Function

The 3D shape determines what a protein does:

  • Enzymes have pockets that fit specific molecules

  • Antibodies have shapes that recognize specific targets

  • Channels have tunnels for specific molecules to pass through

If shape is wrong → protein doesn’t work!

Example: Sickle cell anemia

  • ONE amino acid change in hemoglobin

  • Changes protein shape

  • Causes red blood cells to become sickle-shaped

  • Leads to serious disease

15.5 Techniques in Proteomics

15.5.1 How Scientists Study Proteins

15.5.2 1. Mass Spectrometry (MS)

What it does: Identifies proteins and measures their amounts

How it works:

  1. Break proteins into small pieces (peptides)

  2. Ionize the peptides (give them a charge)

  3. Measure their mass-to-charge ratio

  4. Computer identifies which protein it came from

Like:

  • Weighing puzzle pieces to figure out which puzzle they’re from

  • Very accurate and sensitive!

Used for:

  • Identifying proteins in samples

  • Measuring protein amounts

  • Finding PTMs

  • Discovering new proteins

15.5.3 2. Protein Microarrays

What they are: Chips with thousands of proteins or antibodies

How they work:

  • Put many different proteins on a chip

  • Add your sample

  • See which proteins interact with what

  • Like a dating app for proteins!

Used for:

  • Finding protein-protein interactions

  • Detecting disease biomarkers

  • Drug discovery

15.5.4 3. Western Blotting

What it does: Detects specific proteins

How it works:

  1. Separate proteins by size (gel electrophoresis)

  2. Transfer to membrane

  3. Use antibodies to detect specific protein

  4. Like using a highlighter to find one word in a book

Used for:

  • Confirming presence of specific protein

  • Measuring protein amount

  • Detecting PTMs

15.5.5 4. X-ray Crystallography & Cryo-EM

What they do: Determine 3D protein structures

X-ray crystallography:

  • Grow protein crystals

  • Shoot X-rays through

  • Calculate 3D structure from diffraction pattern

Cryo-EM (Cryo-electron microscopy):

  • Freeze proteins quickly

  • Image with electron microscope

  • Reconstruct 3D structure

  • Nobel Prize 2017!

Used for:

  • Understanding how proteins work

  • Drug design (fitting drugs into protein pockets)

  • Seeing proteins in action

15.5.6 5. Yeast Two-Hybrid

What it does: Finds protein-protein interactions

How it works:

  • Use yeast cells as test tubes

  • If two proteins interact, yeast survives

  • If they don’t interact, yeast dies

  • Like a matchmaking test!

15.6 Applications of Proteomics

15.6.1 Medicine

Disease Diagnosis:

  • Protein biomarkers in blood/urine

  • Early cancer detection

  • Monitoring disease progression

Personalized Medicine:

  • Protein profiles guide treatment

  • Predict drug response

  • Tailored therapies

Drug Discovery:

  • Find new drug targets

  • Design drugs to fit protein structures

  • Understand side effects

15.6.2 Biotechnology

Industrial Enzymes:

  • Laundry detergents

  • Food processing

  • Biofuels

Therapeutic Proteins:

  • Insulin for diabetes

  • Antibodies for cancer

  • Growth factors for wounds

15.6.3 Research

Understanding Life:

  • How cells work

  • Cell signaling pathways

  • Development and differentiation

Evolution:

  • Compare proteins across species

  • Understand evolutionary relationships

  • See how proteins evolved

15.7 The Future of Proteomics

15.7.1 Emerging Technologies

Single-Cell Proteomics:

  • Study proteins in individual cells

  • See cell-to-cell variation

  • Understand rare cell types

Spatial Proteomics:

  • See where proteins are in tissues

  • 3D protein maps

  • Understand protein location and function

Integrative Multi-Omics:

  • Combine genomics + transcriptomics + proteomics

  • Complete picture of cell state

  • Systems biology approach

15.8 Key Takeaways

  • Proteomics = Large-scale study of all proteins

  • Proteome is more complex than transcriptome due to PTMs

  • Post-translational modifications (PTMs) add diversity:

    • Phosphorylation, methylation, acetylation

    • Ubiquitination, glycosylation, cleavage

    • Create over 1 million protein forms from 20,000 genes

  • Protein structure has 4 levels (1°, 2°, 3°, 4°)

  • Structure determines function

  • Proteomics techniques:

    • Mass spectrometry (identify & quantify)

    • Protein microarrays (interactions)

    • Western blotting (detect specific proteins)

    • X-ray crystallography & Cryo-EM (3D structures)

  • Applications: Medicine, drug discovery, biotechnology, research

  • Proteins are the actual workers—understanding them is crucial for understanding life!

Sources: Information adapted from Technology Networks, Abcam, PMC, and proteomics research literature.