15.4 Post-translational modifications

Post-translational Modifications and Protein Function

Proteins don't become fully functional the moment they roll off the ribosome. After synthesis, they undergo chemical changes called post-translational modifications (PTMs) that alter their structure, function, stability, and localization. PTMs let cells fine-tune protein activity quickly, without having to transcribe and translate entirely new proteins every time conditions change.

Equally important is protein folding: a protein's 3D shape determines what it can do, and specialized proteins called chaperones ensure that folding happens correctly. When PTMs or folding go wrong, the consequences range from lost function to serious disease.

Types of Post-translational Modifications

Phosphorylation is the addition of a phosphate group to a protein, carried out by enzymes called kinases. This is one of the most common PTMs and is reversible: phosphatases remove the phosphate group. Adding or removing a phosphate induces a conformational change that can switch a protein "on" or "off."

• Central to cell signaling (e.g., the insulin signaling pathway), metabolism (e.g., regulation of glycogen synthase), and cell cycle control (e.g., cyclin-dependent kinases activating cell division checkpoints)

Glycosylation is the attachment of sugar molecules to a protein. There are two main types:

1. N-linked glycosylation attaches sugars to asparagine residues and occurs in the ER.
2. O-linked glycosylation attaches sugars to serine or threonine residues and occurs in the Golgi apparatus.

Glycosylation serves as a quality-control checkpoint for protein folding, protects proteins from degradation, and influences cell-cell interactions (for example, cell adhesion molecules on the cell surface rely on proper glycosylation to bind their partners).

Ubiquitination is the covalent attachment of a small protein called ubiquitin to a target protein. Its best-known role is tagging proteins for destruction by the proteasome, but ubiquitination also regulates protein localization (e.g., nuclear transport), activity (e.g., transcription factor regulation), and signaling.

Ubiquitination requires a three-enzyme cascade:

1. E1 (activating enzyme) activates ubiquitin in an ATP-dependent step.
2. E2 (conjugating enzyme) carries the activated ubiquitin.
3. E3 (ligase enzyme) recognizes the specific substrate and transfers ubiquitin to it. Substrate specificity comes mainly from the E3.

Protein Folding and Chaperones

How Chaperones Work

Chaperones are proteins that help other proteins fold into their correct 3D conformations. Without them, newly synthesized polypeptides or stress-damaged proteins tend to misfold and clump together into aggregates.

Key families of chaperones include the heat shock proteins (Hsp70, Hsp90, Hsp60) and chaperonins like the bacterial GroEL/GroES complex. They assist folding through several mechanisms:

1. Shielding partially folded proteins from the crowded cytoplasm to prevent aggregation.
2. Providing an isolated folding chamber (the GroEL/GroES complex encapsulates a single polypeptide, giving it a protected space to fold).
3. Refolding proteins that have been denatured by heat shock or oxidative stress.
4. Routing hopeless cases for degradation: the co-chaperone CHIP has ubiquitin ligase activity and can tag terminally misfolded proteins for proteasomal destruction.

Consequences of Improper Modifications

When PTMs go wrong or chaperone systems fail, proteins malfunction, and disease often follows.

• Hyperphosphorylation of tau is a hallmark of Alzheimer's disease. Excess phosphate groups cause tau to detach from microtubules and aggregate into neurofibrillary tangles, disrupting neuronal function.
• Abnormal glycosylation is common in cancer cells. Altered sugar patterns on integrins, matrix metalloproteinases, and selectins promote changes in cell adhesion, tissue invasion, and metastasis.
• Impaired ubiquitination leads to accumulation of misfolded proteins. In Parkinson's disease, mutations in the E3 ubiquitin ligase Parkin prevent proper clearance of damaged proteins and cause mitochondrial dysfunction.

Because PTMs are so central to disease, they're also therapeutic targets. Kinase inhibitors like imatinib treat chronic myeloid leukemia by blocking an overactive kinase. Proteasome inhibitors like bortezomib treat multiple myeloma by disrupting protein degradation in cancer cells. Hsp90 inhibitors are being explored in cancer therapy to destabilize oncogenic proteins that depend on chaperone support.