CLINICAL CORRELATIONS PROTEIN MISFOLDING CLINICAL CORRELATIONS

Protein misfolding Every protein must fold to achieve its normal conformation and function. Abnormal folding of proteins leads to a number of diseases in humans.

Role of chaperons in protein folding: Chaperons are a specialized group of proteins, required for the proper folding of many species of proteins. They also known as “heat shock” proteins. The interact with polypeptide at various stages during the folding process.

• Folding means arriving at the right combinations of angles for every amino acid residue in the sequence. • Proteins fold on a defined pathway (or a small number of alternative pathways); they don't randomly search all possible conformations until they arrive at the most stable (lowest free energy) structure. • Proteins that don't (re)fold on their own, need "molecular chaperones" (which are also proteins) to keep them from slipping off the folding pathway or to help them to get back on it. –Some chaperones require ATP to carry out their function.

MANY DISEASES ARE THE RESULT OF DEFECTS IN PROTEIN FOLDING. • Examples: – CYSTIC FIBROSIS involves misfolding and resulting in lack of a protein involved in Cl– transport across membranes. – MANY NEURODEGENERATIVE DISORDERS INVOLVE ABNORMAL PROTEIN AGGREGATION. • PRION DISEASES (e.g., CJD, Creutzfeldt-Jakob disease) = spongiform encephalopathies (also includes “mad cow” disease, scrapie in sheep, etc.) • Alzheimer disease • Parkinson Disease • Huntington Disease • PARTLY FOLDED OR MISFOLDED POLYPEPTIDES OR FRAGMENTS may sometimes associate with similar chains to form aggregates. • Partial unfolding of correctly folded proteins may also lead to aggregation. • Aggregates vary in size from soluble dimers and trimers up to insoluble fibrillar structures (amyloid). AMYLOIDOSIS

OTHER DISEASES INVOLVING ABNORMAL FOLDING/AGGREGATION OF OTHER PROTEINS • HUNTINGTON DISEASE (a "polyglutamine disease", trinucleotide expansion disorder) – Hereditary: autosomal dominant allele, caused by mutation that‘s an expansion (abnormal lengthening) of repeated sequence CAGCAGCAG… in gene for the protein huntingtin, encoding polyGln sequences in the protein. – Huntington disease affects about 1 in 10,000 people in general population -- late onset (middle age), but longer triplet expansions seem to cause earlier onset of more rapidly progressing disease (due to the more highly aggregation-prone mutant protein with longer polyGln sequences) – Protein aggregates accumulate in nuclei and cytoplasm of neurons in striatum region of the brain, which is required for coordinated movements, ultimately killing the neurons. – Disease leads to rigidity and dementia, always fatal. – Other trinucleotide expansion diseases, e.g., spinocerebellar ataxias.

CYSTIC FIBROSIS • defective protein = CFTR (Cystic Fibrosis Transmembrane conductance Regulator) • LACK of normal protein, not the abnormal protein itself, causes disease. • Normal protein is a membrane protein (an ATP-regulated chloride channel) in plasma membranes of epithelial cells, that pumps Cl– ions OUT of cells • Defective (mutant) protein doesn’t fold properly. • Folding intermediates don't dissociate from chaperones, preventing CFTR from insertion into membrane. • High intracellular Cl– concentration makes cells take up H2O from surrounding mucus by osmosis. • Thick mucus accumulates in lungs and other tissues, and its presence in lungs causes difficulty breathing and makes affected individuals very subject to infections like pneumonia.

HUMAN NEURODEGENERATIVE DISORDERS e.g., PRION DISEASES like Creutzfeldt-Jakob Disease (CJD), Alzheimer Disease, Parkinson Disease, Huntington Disease • formation of protein aggregates, amyloid plaques/tangles, lesions in the brain (Other amyloid-forming diseases affect other organs, e.g., liver or heart). • Fibrils resulting from aggregation of different proteins (different diseases) have common structural feature: central core of β sheets known as a "cross-beta" structure.

SPONGIFORM ENCEPHALOPATHIES e.g., scrapie (sheep), kuru (humans), bovine spongiform encephalopathy (BSE, "mad cow disease"), scrapie (sheep), kuru (humans), Hypothesis: misfolded proteins in all kinds of cells are always forming cross-beta structures, but most cell types have adequate degradative machinery to clean up the "garbage" so it doesn't accumulate. Maybe neurons lack adequate molecular machinery to dispose of them. Abnormal protein can be acquired by infection, or by inheritance (dominant), or spontaneously ("sporadic" -- unknown cause).

(SPONGIFORM ENCEPHALOPATHIES, continued) • A tiny bit of ABNORMALLY folded protein, PrPSC, can aggregate to form “cross-beta” structures, and small aggregates ("nuclei") can shift folding of normal proteins to join the aggregation process and adopt the abnormal conformation • In a sort of cascading "domino effect", a high concentration of abnormally folded protein (PrPSC) forms amyloid plaques (large insoluble fibrous aggregates with β conformation prevalent) in brain. • These cross-beta structures fortunately don't form very fast, but if concentration of the aggregation-prone intermediates increases, rate of cross-beta formation can increase dramatically.

Creutzfeldt-Jacob or prion disease: Prion protein is present in normal brain tissue.In diseased brains, the same protein is misfolded. It, therefore, forms insoluble fibrous aggregates that damage brain cells. • Fatal, neurodegenerative diseases, with characteristic "holes" appearing in brain ("sponge"-like appearance) • Infectious, but causative agent is an abnormal protein, a "prion” ("proteinaceous infectious only" protein, PrP) (Stanley Prusiner, Nobel Prize in Physiology/Medicine 1997). 1. Transmissible agent: various sized aggregates of a specific protein 2. Aggregates are resistant to treatment by most protein-degrading enzymes. 3. Protein is largely or completely derived from a cellular protein, PrP, (PrPC) normally present in brain. 4. PrPC has a lot of α-helical conformation; abnormal conformation, PrPSC, has much more β conformation, that tends to aggregate with other PRP molecules.

Alzheimer Disease : β amyloid protein is a misfolded protein. Protein misfolding Alzheimer Disease : β amyloid protein is a misfolded protein. It forms fibrous deposits or plaques in the brains of Alzheimer’s patients. • Symptoms: memory loss, dementia, impairment in other forms of cognition and behavior • Not transmissible between individuals • Intracellular aggregates (fibrillar tangles) of protein called tau • Extracellular plaques contain aggregates of β-amyloid peptides (Aβ):40-42-residue segments derived by proteolytic cleavage of a much larger protein (amyloid precursor protein, APP) attached to plasma membrane of neurons (function unknown)

OTHER DISEASES INVOLVING ABNORMAL FOLDING/AGGREGATION OF OTHER PROTEINS • PARKINSON DISEASE – Protein that forms fibrils is α-synuclein. – Lesions ("Lewy bodies") form in cytosol of dopaminergic neurons in substantia negra of brain, accompanied by muscle rigidity and resting tremor.

In the brain, alpha-synuclein is found mainly at the tips of nerve cells (neurons) in specialized structures called presynaptic terminals. Within these structures, alpha-synuclein interacts with phospholipids and proteins. Presynaptic terminals release chemical messengers, called neurotransmitters, from compartments known as synaptic vesicles. The release of neurotransmitters relays signals between neurons and is critical for normal brain function. Although the function of alpha-synuclein is not well understood, studies suggest that it plays a role in maintaining a supply of synaptic vesicles in presynaptic terminals by clustering synaptic vesicles. It may also help regulate the release of dopamine, a type of neurotransmitter that is critical for controlling the start and stop of voluntary and involuntary movements