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Protein Folding in the Cell 1

BIOC 212

Winter 2013

J ason C. Young

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Lecture Topics

Protein Folding

1. Elements of protein structure

2. Molecular chaperones3. Multi-chaperone systems

4. Modifications and Degradation

Membranes

1. Membrane lipids2. Membrane proteins

3. Targeting to endoplasmic reticulum (ER)

4. Sorting and degradation in ER

Intracellular Traffic1. Vesicle formation

2. Vesicle targeting and fusion

3. Lysosome and Nucleus

4. Mitochondria

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I am a protein. All living organisms need me to function. A basicbuilding block of the human body, Im made from amino acids found

in ribosomes. Proteins give energy to everything from flowers andbutterflies to heroes who turn in Communists. I...am a protein.

Jack Donaghy, 30 Rock Chain Reaction of Mental Anguish

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The Cell

David Goodsell, Scripps

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Cellular Proteins

Proteins are the main functional components in cells

Genes and mRNA are linear

Proteins are made as linear polypeptides by cytosolic ribosomes,but fold into 3-dimensional conformations

Folding provides physical stability and functional surfaces The sequence of a protein determines its structure, function and

localization

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Amino Acids

20 different amino acids

Side chains have different chemical characteristics:

hydrophobic, polar or charged (acidic or basic)

small or large

covalently linked into polypeptides

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Polypeptides

Peptide bonds in the backbone are uncharged but polar

Charge and hydrophobicity of a polypeptide is determined by the

side chains

Both side chains and backbone can form non-covalent contacts withother amino acids

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Polypeptide Backbone

The peptide bond is planar and cannot rotate

Rotation around the bonds to the central carbon (C) is possible

Therefore, the polypeptide backbone has limited freedom of rotation

Some rotation angles between amino acids (residues) in apolypeptide are preferred

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Non-Covalent Bonds

Interactions between residues of a polypeptide stabilize structure

hydrophobic interactions (exclusion of water)

hydrogen bonds

van der Waals interactions (transient dipoles between all atoms)

ionic bonds

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Protein Folding

Folding is driven by hydrophobic interactions

other non-covalent interactions and rigidity constraints contribute

to the structure

Polar side chains usually form outer surface

Native State is the most stable conformation

Proteins with similar sequences usually have similar native states,and may have similar functions

native state

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Disulfide Bonds

Secretory proteins often have covalent disulfide bonds betweencysteine side chains

extracellular proteins, inside secretory organelles

disulfides reinforce structure

Cytosolic proteins do not have disulfide bonds

cytosol, nucleus, mitochondria

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Importance for Folding

strong hydrophobic interactions are important for protein structure,and also for membrane formation

hydrophobic interactions very many, strong

hydrogen bonds many, strong

Van der Waals interactions many, weak

ionic bonds few, strong

disulfide bonds few, very strong

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Units of Protein Structure

Primary structure amino acid sequence

Secondary structure local conformation patterns

stretches of polypeptide can have regular arrangements of thepolypeptide backbone and position of side chains

common structures are -helices and -sheets loops have no regular secondary structure and can be flexible

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Alpha-Helix

-helix:

backbone is coiled

hydrogen bonds between each turn of helix backbone

side chains point outwards

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Beta-Sheets

-strands

backbone is extended almost straight

several strands pack sideways into a -sheet hydrogen bonds between the backbone strands

side chains on alternate sides

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Tertiary Structure

Tertiary structure complete three-dimensional arrangement of thepolypeptide

secondary structure elements are packed against each other toform the tertiary structure

hydrophobic contacts between secondary elements

long-range contacts between residues that are far apart in theprimary sequence

loops

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Units of Protein Structure 4

Quaternary structure: the assembly of multiple polypeptides(subunits) into a final protein

interactions between subunits are very stable monomer: single polypeptide with no quaternary structure

dimer: two polypeptide subunits

trimer, tetramer, 5-mer, 6-mer etc. oligomer: many subunits

haemoglobin tetramer

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Domains

A domain is an independently folded unit within a protein

Proteins can have one or multiple domains

Different domains in a protein often have different functions

Hsp70 ATPase

domain

ribbon diagram(polypeptidebackbone only)

space-filling model(all atoms)

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Polypeptide Length

Most polypeptides are 100 to 800 amino acids long, or from 12 kDato 90 kDa molecular weight

Domains are usually 50 to 200 amino acids long Long proteins have multiple domains

0

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length (amino acids)

number

ofproteins

proteins encoded byhuman chromosome 1

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Protein Surface

The sequence of a protein determines its functional surface andinteractions with ligands

Specificity of binding can be narrow (few molecules recognized) orbroad (many different molecules)

Catalysis: some proteins are enzymes which increase rate ofcovalent chemical reactions

Allostery: conformational changes can change binding surface

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Modular Domains

Some types of domains are found in many different proteins

Many modular domains form reversible, non-covalent contacts with

specific features on other molecules other proteins (different from quaternary structure)

certain lipids or carbohydrates

allow regulation of function

examples of proteins withmodular J domains (green oval)

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Some amino acid side chainsare chemically similar to each

other

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Protein Families

Proteins or domains in a family have similar sequences andstructures; they usually have related functional mechanisms

Similarity (homology) indicates evolutionary conservation Divergent sequences have no similarity and different structures, but

may have still have related functions

homologous

human Hsp70ATPase domain

E. coli Hsp70ATPase domain

E. coli arsenitetransporter ATPasesubunit

not homologous

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Which amino acid side chains can form these interactions?

can the polypeptide backbone form any of these interactions?

hydrophobic interactions

hydrogen bonds

Van der Waals interactions

ionic bonds

disulfide bonds

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End of 1

Molecular Biology of the Cell, Alberts et al. 4th or 5th Ed.

Ch. 3, protein structure, protein function

Dobson (2003) Protein Folding and Misfolding. Nature 426, 884-890.