Chapter 6 Protein Structure and Folding

1. Secondary Structure 2. Tertiary Structure 3. Quaternary Structure and Symmetry 4. Protein Stability 5. Protein Folding

Myoglobin Introduction

1. Proteins were long thought to be colloids of random structure 2. 1934, crystal of pepsin in X-ray beam produces discrete diffraction pattern -> atoms are ordered 3. 1958 first X-ray structure solved, sperm whale myoglobin, no structural regularity observed 4. Today, approx 50’000 structures solved => remarkable degree of structural regularity observed Hierarchy of Structural Layers

1. Primary structure: amino acid sequence 2. Secondary structure: local arrangement of peptide backbone 3. Tertiary structure: three dimensional arrangement of all atoms, peptide backbone and amino acid side chains 4. Quaternary structure: spatial arrangement of subunits

1) Secondary Structure

A) The planar peptide group limits polypeptide conformations The peptide group ha a rigid, planar structure as a consequence of resonance interactions that give the peptide bond ~40% double bond character The trans peptide group The peptide group assumes the trans conformation 8 kJ/mol mire stable than cis Except Pro, followed by cis in 10% Torsion angles between peptide groups describe polypeptide chain conformations

The backbone is a chain of planar peptide groups The conformation of the backbone can be described by the torsion angles (dihedral angles, rotation angles) around the Cα-N (Φ) and the Cα-C bond (Ψ)

Defined as 180° when extended (as shown)

+ = clockwise, seen from Cα Not all combinations of torsion angles are possible, some result in collision of N-H with O=C or R The Ramachandran diagram indicates allowed conformations of polypeptides

• Pro limits Φ to -60°

• Gly allows more freedom B) The most common regular secondary structures are the α Helix and the β Sheet

• Two structures are widespread: α Helix and β Sheet = regular secondary structures • The α helix is a coil: structure with allowed angles and stabilized by hydrogen bonding

• Linus Pauling, 1951, model building

• Right-handed turn

• Pitch 3.6 Aa/turn = 5.4 Å

• Average length 12 Aa = 3 turns ~18 Å

• Bonding of C=O with N-H at n+4

• R point outwards, downwards

β sheets are formed from extended chains • like α helix, β sheets are stabilized by hydrogen bonding • Bonding occurs between neighboring chains rather than within the structure as is the case with α helix • Parallel and antiparallel

Pleated appearance of a β sheet • From 2 to 22 strands • Average 6 strands in protein • Exhibit right-handed twist

7-stranded Bovine carboxypeptidase A

Coils Helices β-sheet Turns connect some units of secondary structure

• topology = connectivity of β sheets can be complex or made by a simple turn/loop • reverse turns or β bends, occur at the protein surface, connect β sheets Reverse turns in polypeptide chains

• Require 4 Aa • 180° flip of Aa 2 C) Fibrous proteins have repeating secondary structure

Historical classification of proteins as globular of fibrous

Fibrous proteins often have protective, connective, or supportive role in living organisms examples: keratin and collagen α keratin is a coiled coil

Keratin, in all higher vertebrates - horny outer epidermal layer - hair, horn, nails, feathers - α in mammals - β in birds and reptiles - Humans, over 50 α genes - tissue specific expression

Forms coiled coil structure - helix pitch 5.1 Å (rather than 5.4 Å as in α-helix) - Parallel N->C, 81 residues long Coiled coil has a pseudo-repeating structure: a-b-c-d-e-f-g in which residues a and d are predominately nonpolar.

Helical wheel representation Higher order α keratin structure -> Macrofibril

-> single hair α keratin

α keratin is rich in Cys residues, form disulfide bonds that crosslink adjacent chains

α keratins are classified “hard” and “soft” depending on whether they have high or low sulfur content

Hard keratins in hair, horn, and nails are less pliable than soft keratins such as in skin and callus (hornhaut)

Reductive cleavage of disulfide bonds results in “curled hairs” Collagen is a Triple Helix

Collagen occurs in all multicellular organism, - Is the most abundant vertebrate protein - Strong insoluble fibers - Stress bearing component of connective tissues (bone, teeth, cartilage, tendon)

One collagen molecule consists of three polypeptide chains - Mammals have at least 33 genetically distinct chains - Assembled in 20 different collagen varieties - One of the most common is Type I collagen:

- two α1(I) chains and one α2(I) chain Collagen has a distinctive amino acid composition: - 1/3 of its residues is Gly - 15 to 30% are Pro and 4-hydroxyprolyl (Hyp)

These non-standard amino acids are formed after collagen is synthesized - Pro - > Hyp conversion by prolyl hydroxylase, Vitamin C-dependent (ascorbic acid) Collagen Diseases Scurvy (Skorbut) results from Vitamin C-deficiency, - Lack of newly synthesized functional collagen, - Skin lesion, fragile blood vessels, poor wound healing - Sailors on long trips, lack of fresh food - Captain James Cook “limely” introduced limes to diet

Several rare genetic collagen disorders - tend to be dominant The Collagen Triple Helix

Collagen amino acid sequence consists of a repeating triplet: Gly-X-Y over some 1000 residues, X often Pro, Y often Hyp - Pro prevents formation of α helix - Collagen forms left-handed superhelix with ~3 amino acids per turn - Three parallel chains wind around each other to form right-handed triple-helical structure - Every third residue of each chain passes through the center if the triple helix, only Gly can pass - Three chains are staggered, Gly at each level - Inter-chain hydrogen bonding Structure of collagen model peptide

Hydrogen bonding from Gly N- to Pro O-atoms of adjacent chain Collagen Crosslinking

Collagens well-packed, rigid, triple-helical structure is responsible for its characteristic tensile strength

Collagens assemble to form loose networks or thick fibrils arranged in bundles or sheets, depending on the tissue

Collagen is internally covalently cross linked which makes it insoluble, not by Cys but by Lys and His reactions

Lysyl oxidase reaction:

D) Most proteins include nonrepetitive structures Majority of proteins are globular and may contain several types of regular secondary structures, including α helices and β sheets and coils (≠ random coils, which are completely unstructured)

The sequence affects the secondary structure: - Pro is helix breaking, induces kink in helix and β sheet - Large amino acids such as Tyr and Ile induce steric clashes - Peter Chou and Gerald Fasman revealed the propensity of amino acids to form helical or sheet structures -> Chou-Fasman prediction

2) Tertiary Structure

Tertiary structure of a protein describes the folding of its secondary structural elements and specifies the position of every atom in the protein

This information is deposited in protein structure database (pdb)

Experimentally determined by X-ray crystallography or NMR Protein crystals A) Most proteins structures are determined by X- ray crystallography or nuclear magnetic resonance X-Ray crystallography: technique that directly images molecules - X-ray wave length is short, ~1.5 Å, equivalent to distance of atoms (visible light 4000 Å) - Crystal: repetitive arrangement of the same structure => diffraction pattern (darkness of spot is function of crystals electron density) - X-ray interact with electrons (not with nuclei) -> X-ray structure is thus an electron density map of a given protein -> represents contours of atoms A thin section through a 1.5 Å resolution electron density map of a protein that is contoured in three dimensions Most protein crystal structures exhibit less than atomic resolution Crystal is build up by repeating units, containing protein in native conformation, - highly hydrated (40-60% water) - soft, jellylike consistency, unlike NaCl crystals - molecules are slightly disordered and display Brownian motion -> this determines the resolution limit of a given protein crystal (typical 1.5 – 3 Å) - inability to crystallize a protein to form crystals of sufficiently high resolution is a major limiting factor in structure determination Electron density maps of diketopiperazine at different resolution levels

- Electron density map alone is not sufficient to determine the structure if the protein, - Amino acid sequence is also required - Computerized fitting algorithm of atoms into the experimentally determined electron density map results in protein structure determination of up to 0.1 Å resolution Most crystallized proteins maintain their native conformations

Key question: does the structure of protein in a crystal accurately reflect the structure of the protein in solution, where it normally functions ?

1. The protein in the crystal is hydrated like it is in solution 2. X-ray structure is similar to NMR structure, which is determined from proteins that are in solution 3. Many enzymes remain catalytically active in the crytsal Protein structures can be determined by NMR Nuclear magnetic resonance, NMR, an atom nucleus resonates if a magnetic field is applied. This resonance is sensitive to the electronic environment of the nucleus and its interaction with nearby nuclei - Developed since 1980, Kurt Wüthrich (ETH-Z), to determine protein structures in solution - Because there are many nuclei in a protein that would crowd in a conventional one-dimensional NMR -> two- dimensional (2D) NMR was developed to measure atomic distances of chemically linked atoms (COSY) or of spatially close atoms (NOESY) - Size limit of about 40 kD, may reach 100 kD soon - Dynamic, can follow protein motion or folding

B) Side chain location varies with polarity Since Kendrew solved the first protein structure, nearly 50’000 protein structures have been reported No two are the same, but they exhibit some remarkable consistency: globular structures lack the repeating sequences that support the conformation of fibrous proteins Amino acid side chains in globular proteins are distributed according to their polarity: 1. Nonpolar residues Val, Leu, Ile, Met and Phe occur mostly in the interior of a protein, excluded from the contact with water, hydrophobic core, compact packing (no empty room) 2. Charged amino acids Arg, His, Lys, Asp, Glu are located on the surface, never in hydrophobic core 3. Uncharged polar groups Ser, Thr, Asn, Gln, Tyr are usually on the surface but are also found inside but then are hydrogen bonded Side chain locations in an α helix and a β sheet

polar nonpolar

Surface of protein Side chain distribution in horse heart cytochrome c

Hydrophilic amino acids Hydrophobic amino acids C) Tertiary structures contain combinations of secondary structure

Globular proteins are build from combinations of secondary structure elements

These combinations of secondary structure elements form protein motifs = supersecondary structures 1. Most common is βαβ motif, α helix connects two parallel strands of a β sheet 2. Equally common is a β hairpin, antiparallel strands connected by tight reverse turn 3. αα motif, two successive antiparallel α helices packed against each other 4. Greek key motif, β hairpin is folded over to form 4- stranded antiparallel β sheet Most proteins can be classified as α, β, or α/β

Secondary structural elements occur in globular proteins in varying proportions

- E. coli cytochrome b562 for example consists only of α helices => α protein - Immunoglobulins contain the immunoglobulin fold => β proteins, contain large proportion of β sheets - Most proteins, including lactate dehydrogenase and carboxypeptidase A are α/β proteins (average ~31% α helix, 28% β sheet) - Further subdivision of proteins by their topology: that is connection of secondary structural elements Selection of protein structures

Cytochrome b562 with heme Immunoglobulin Lactate fragment dehydrogenase Structures of β barrels Human retinol Peptide Triose binding protein amidase F phosphate isomerase Large polypeptides form domains

Polypeptides of more than 200 amino acids usually fold into more than one domain in eukaryotes, prokaryotes can only fold mono-domain proteins -> bi or multilobal appearance

Most domains consist of 40 to 200 Aa, average diameter of ~25 Å

Many domains are structurally independent units that have the characteristic of globular proteins

Individual domains often have specific function, i.e. binding of the dinucleotide NAD+ by nucleotide binding site:

Rossmann fold: 2 (βαβαβ) Glyceraldehyde-3-phosphate dehydrogenase

2 globular domains Dinucleotide binding in N-term domain Rossmann fold D) Structure is conserved more than sequence

- Grouping structures into families of high similarity, - 50’000 structures define 1’400 protein domain families - 200 different folding patterns account for about half of all known structures - the protein domain is the evolutionary unit, not its sequence - comparison of c-type cytochromes E) Structural bioinformatics provides tools for storing, visualizing, and comparing protein structural information

Structural data obtained by X-ray or NMR describing the room coordinates of atoms is deposited into database, similar to sequence information of DNA or proteins

Bioinformatics, structural bioinformatics takes advantage of this information to address biological questions

Major structural database: Protein Data Bank (PDB), each structure is assigned a unique identifier (PDBid), i.e., sperm whale myoglobin is 1MBO

Molecular graphics program interactively show macromolecules in three dimensions

- Jmol is a Web browser-based application that allows you to directly visualized structures in PDB

- Example potassium channel (KCSA) http://www.pdb.org/pdb/explore/jmol.do? structureId=1F6G

- Swiss PDB viewer = Deep View allows protein modeling and superimposition of two structures Structure comparisons reveal evolutionary relationships - Since evolution tends to conserve structure rather than sequence, programs have been developed to search for structurally related protein

- CATH, classifies proteins in a four level hierarchy: 1. Class: mainly α / β structure 2. Architecture: gross arrangement of secondary structure 3. Topology: shape of protein domains and interconnectivity 4. Homologous superfamily, group of common ancsestor - CE (combinatorial extension of the optimal path) finds all proteins in PDB that can be structurally aligned with the query structure - FSSP (Family of Structurally Similar Proteins) - SCOP (Structural Classification of Proteins) - VAST (Vector Alignment Search Tool) 3) Quaternary Structure and Symmetry - Many proteins, particularly those of > 100 kD consist of more than one polypeptide chain. Multi-subunits associate into defined structure = quaternary structure

- For example collagen, assembly of multiple subunits is easier than synthesizing one very large polypeptide chain…. - Site of synthesis can differ from site of assembly - Damaged components can be replaced - Less genetic information required to encode self-assembling subunits - Multi-subunit enzymes have multiple catalytic sites that can be co-regulated Subunits usually associate noncovalently

- Multi-subunit protein may consist of identical or nonidentical subunits (homo-, hetero-oligomeric) - oligomers, protomers - Example: hemoglobin is a dimer of αβ protomers

- Contact region between subunits resembles the interior of a single subunit protein: closely packed nonpolar residues, hydrogen bonding, interchain disulfide bridges, but generally less hydrophobic than the hydrophobic core of a single subunit protein (they are synthesized as monomers and need to be soluble each one before assembly…) Quaternary structure of hemoglobin

heme Subunits are symmetrically arranged - In the majority of oligomeric proteins, the subunits are symmetrically arranged - That is: protomers occupy geometrically equivalent positions

- No inversion or mirror symmetry because this would require D-amino acids

- Thus proteins can have only rotational symmetry

- Simples case, cyclic symmetry, single axis of rotation (2-,3-,

4-,n-fold). C2 is most common

- Dihedral symmetry (Dn): n-fold rotational axis intersects with a twofold rotational axis. D2 most common - Tetrahedron, cube, and icosahedron, for example spherical viruses Symmetries of oligomeric proteins

Rotational symmetry Symmetries of oligomeric proteins

Dihedral symmetry Symmetries of oligomeric proteins 4) Protein Stability

- Native proteins are only marginally stable under physiological conditions -> high turnover

- Free energy required for denaturation is ~0.4 kJ/mol/ residue -> fully folded 100 residue protein is ~40 kJ/mol more stable than its unfolded form = energy of 2 hydrogen bonds

- But energy of all noncovalent interactions within a protein is in the order of thousands of kJ/mol

=> native structure results from a delicate balance of powerful counteracting forces A) Proteins are stabilized by several forces - Protein structures are governed mainly by hydrophobic effects and to lesser extend by interactions between polar residues

- The hydrophobic effect causes nonpolar substances to minimize their contact with water (degree of order, entropy, of water is decreased because water has not to form “cages” around the hydrophobic groups)

- Relative hydropathy of residues: energy required to solubilize a given amino acid in water

Hyrdopathic index plot for bovine chymotrypsinogen

interior

exterior

Sum of hydropathie of 9 consecutive residues is plotted Electrostatic interactions contribute to protein stability - Relatively week van der Waals forces are nevertheless important to stabilize the protein in its native state

- But hydrogen bonds, which are a central feature of protein structures, particularly secondary structures, make only a minor contribution to the overall stability of a protein

- Because of extensive hydrogen bonding of surface residues to water, difference between native and unfolded energy of hydrogen bonding is ~-2 to 8 kJ/mol

- Ion pairing / salt bridges, i.e. between Lys+ and Asp- 75% of ionized residues are paired, mostly on surface, but they have only a small stabilizing effect (paid by loss of entropy and loss of solvation free energy) => poorly conserved Examples of ion pairs in myoglobin

- Oppositely charged side chains from groups that are far apart in sequence closely approach each other through the formation of ion pairs Disulfide bonds cross-link extracellular proteins - Intrachain or inter-chain disulfide bonds form during folding of the protein

- While they are not necessary for the structure/function of most proteins, they “lock in” a particular backbone fold

-Disulfide bonds are rarely in cytoplasmatic protein because the cytosol is a reducing environment that would cleave the disulfide bridges

- Most disulfide bridges occur in secreted proteins, i.e. they are stable in the more oxidizing extracellular environment Metal ions stabilize some small domains

- Metal ions, bound within a protein, can also serve to stabilize the protein structure, through internal cross-linking - For example zing fingers are motifs that frequently occur in DNA binding proteins - Zn2+ is tetrahedrally coordinated by side chain Cys, His and occasionally Aps and Glu - Zn2+ allows short stretches of polypeptides, 25-60 residues, to fold into stable units - Zinc fingers are not stable in the absence of Zn2+ - Zn2+ is stable and not oxidized/reduced (unlike Cu, or Fe) A zing finger motif

Coordinated by Cys and His Proteins are dynamic structures

- Plethora of forces that act to stabilize a protein leave room for mobility/movement of structural elements - Proteins are flexible and rapidly fluctuating molecules - Mobility is important for protein function - Conformational flexibility = breathing Molecular dynamics of myoglobulin

Snapshots at intervals of 5 x 10-12 sec

Hem B) Proteins can undergo denaturation and renaturation

- Low conformational stability of native proteins makes them susceptible to denaturation: through alterations in the balance of weak bonding forces - Denaturation by: - Heat, cooperative melting of the structure - pH variation, alters ionization state of side chains - Detergents, break hydrophobic interactions - Chaotropic agents, guanidium ion, urea (5-10 M). Small organic molecules that increase the solubility of nonpolar substances in water, disrupt hydrophobic interactions

Many denatured proteins can be renatured

- 1957, Christian Anfinson could denature and renature ribonuclease A (RNase A), 124 Aa single chain protein - Denaturation in 8 M urea - Renaturation by dialysis, full enzymatic activity ⇒ Protein must spontaneously fold into active conformation, including proper re-formation of 4 Cys-bridges ⇒ Even correct Cys bridges form spontaneously: 1/7 x 1/5 x 1/3 x 1/1 = 1/105 (less than 1%) ⇒ The primary structure if the protein dictates its three dimensional conformation Denaturation and renaturation of RNase A Thermostable Proteins - Thermophilic and hyperthermophilic bacteria can grow at near 100°C - Live in hot springs or submarine hydrothermal vents - But have the same metabolic pathways as mesophilic organisms - No striking differences in overall structures - Difference of stability of corresponding thermophilic proteins is ~100 kJ/mol = few noncovalent interactions - Network of salt bridges at surface - Increased hydrophobic contribution, but also only ~0.4 kJ/ mol/residue => low stability is important for function !! Breathing of structure 5) Protein Folding

- Protein folding is directed largely by residues that occupy the interior of the folded protein - How does a protein fold into its three dimensional structure - Does not occur through sampling of all possible conformations ! This would take longer than the universe exists

(n residues -> 2n torsion angles, each has 3 stable conformations ->32n = ~10n possible conformations, 10-13 sec for each conformation -> t = 10n/1013 => for n=100 residues t = 1087 sec, 20 Mia years = 6 1017 sec) A) Proteins follow folding pathways

- Many proteins fold into their native conformation in less than a few seconds - They follow directed pathways, not random - Folding occurs locally by the formation of secondary structures - Followed by a hydrophobic collapse of the structure to adopt a molten globule = has most of the secondary structures formed but not yet the proper tertiary structure - Proteins fold in a hierarchical manner - Cooperative process Energy-entropy diagram for protein folding

Folding funnel

Temporary folding traps Protein structure prediction and protein design

- Sequence of 1 Mio proteins is know, but structure has been determined for only 50’000 - How is the structure encoded in the primary sequence ? -> ab initio prediction of structure - Homology modeling of new sequence against existing structure - Structural genomics, determine X-ray structure for all the representative protein domains in a genome - Chou & Fasman predictions does not take into account the influence of the neighboring residues Protein structure prediction and protein design - Protein design, inverse of structural prediction - Design an amino acid sequence that will form the target structure or even target function - 28 residue peptide that forms ββα structure Protein disulfide isomerase acts during protein folding

- Proteins fold more slowly in vitro than they fold in vivo - This is frequently due to the formation of non- native disulfide bridges which are then slowly exchanged to the native ones - In vivo, disulfide bond formation is catalyzed by and enzyme: protein disulfide isomerase (PDI) - PDI binds a variety of unfolded proteins via a hydrophobic patch to form a mixed disulfide Mechanism of protein disulfide isomerase Mechanism of protein disulfide isomerase B) Molecular chaperones assist protein folding

- Proteins begin to fold as they are synthesized and grow on the ribosome - In vivo, a peptide chain folds in the presence of a very high concentration of other proteins - Molecular chaperones are essential proteins that help to fold newly synthesized or partially unfolded proteins to re-fold correctly - Many molecular chaperones were first described as heat shock proteins (Hsp), their expression is strongly induced upon heat treatment of cells Chaperone activity requires ATP Classes of molecular chaperones in prokaryotes and eukaryotes 1. Hsp70 family, function as monomers with the cochaperone Hsp40, folds newly made proteins

2. Chaperonins, large multisubunit proteins (see below) 3. Hsp90 proteins, folding of proteins in signal transduction such as steroid receptors 4. Trigger factor, associate with ribosome and prevent improper folding of newly made All of them operate by binding to solvent-exposed hydrophobic surfaces and subsequent release All are ATPases The GroEL/ES chaperonin forms closed chambers in which proteins fold The chaperonins in E. coli consist of two types of subunits, GroEL and GroES

Structure: 14 identical 549-residue GroEL subunits arranged in two stacked rings of seven subunits each

Complex is capped at one end by domelike heptameric ring of 97 Aa GroES subunits

Bullet-shaped complex with C7 symmetry Central chamber of ~45 Å in which peptides fold X-ray structure of the GroEL-GroES-(ADP)7 complex GroES (cap)

GroEL (cis)

GroEL (trans) Note the larger size of The cavity formed by the cis ring ATP binding and hydrolysis drive the conformational changes in GroEL/ES

Each GroEL subunit can bind and hydrolyze ATP which induces a conformational change -> movement -> work

All 7 subunits acts in concert to work on the unfolded substrate protein

Exposure (ATP) or hiding (ADP) the hydrophobic patch domain, to allow the protein to refold within an isolated hydrophilic microenvironment

Eukaryotic counterpart: TRiC Reaction cycle of the GroEL/ES chaperonin Some diseases are caused by protein misfolding At least 20 – usually fatal – human diseases are associated with extracellular deposition of normally soluble proteins as insoluble fibrous aggregates = amyloids (starch-like)

Amyloidoses: set of rare inherited diseases in which mutant forms of normally soluble proteins, such as lysozyme or fibrinogen, accumulate as amyloids

Symptoms usually become apparent only later in life (30-70 years) progress over 5-15 years till death

Amyloid-β protein accumulation in Alzheimer’s disease • Alzheimer’s disease is a neurodegenerative condition, mainly in elderly ~10% of >65; 50% >85

• Amyloid plaque in brain tissue, surrounded by dead and dying neurons

• Plaques consist of fibrils of a 40- to 42-residue protein, amyloid-β protein (Aβ)

• Aβ is a fragment from a 770-residue membrane protein, Aβ precursor protein (βPP) whose normal function is unknown Amyloid-β protein accumulation in Alzheimer’s disease (2) • Aβ is excised from βPP in a multistep process through the action of two proteolytic enzymes: β- and γ- secretase • The age-dependence suggest that Aβ deposition is an ongoing process • Rare mutations in βPP result in early onset of the disease • Similar to Down’s syndrome patients (trisomy of Chr 21), which invariably develop Alzheimer’s by their 40th • β- and γ-secretase inhibitors are being developed and tested Brain tissue from an individual with Alzheimer’s disease Prion diseases are infectious

• Scrapie, neurological disorder in sheep and goats, thought to be due to “slow viruses” • Bovine spongiform encephalopathy (BSE, mad cow disease), and kuru (degenerative brain disease in human, cannibalism on Papua New Guineas), Kreutzfeld-Jakob disease (CJD), sporadic (spontaneously arising) • Neurons develop large vacuoles that give brain tissue a spongelike microscopic appearance • All are known as transmissible spongiform encephalopathies (TSEs) Prion diseases are infectious (2) •TSEs are not caused by virus or microorganism •Infectious agent is a protein = prion (proteinaceous infectious particle), TSE = prion diseases •The scrapie prion is named PrP (prion protein), 208 Aa, many hydrophobic residues, aggregates to clusters of rodlike particles that resemble amyloid fibrils •How are prion diseases transmitted ? Scrapie form of PrP, PrPSc is identical to PrPC in sequence but differs in secondary and tertiary structure •PrP Sc catalyzes conformational change of PrPC to become PrPSc which becomes insoluble and forms rodlike particles Electron micrograph of a cluster of partially proteolyzed prion rods

• PrPSc is insoluble and protease resistant • Transmission of BSE to human, new variant CJD in UK (160 cases)

• BSE due to feeding meat-and- bone meal Amyloid fibrils are β sheet structures

• PrPC mostly α helical, PrPSc β sheet structures • β sheet perpendicular to fibril axis • Almost any protein can be induced to form aggregates

PrPC PrPSc Model of an amyloid fibril

• Catalyzed formation from a nucleus/template (rare, slow) • Chain/fibril growth (fast) • β sheet growth mostly stabilized by inter-chain hydogen bonds