Protein Structure
I.
BUILDING BLOCK
A. Proteins are constructed of amino acids
1. Dipeptide
a) Two amino acids linked together
2. Oligopeptide
a) 3 - 9 amino acids linked together
3. Polypeptide
a) Nine or more amino acids linked together
4. Protein
a) May be a polypeptide or an association of several to many polypeptides
(1) Example: Hemoglobin contains four polypeptides and eukaryotic DNA
polymerases contain at least 10 polypeptides
b) A typical protein has between 135 to 635 amino acids
(1) The average molecular weight of an amino acid is 110
(2) Therefore, a typical protein has a molecular weight between 15,000 to 70,000
B. Basic structure
1. All amino acids have an  carbon that forms four bonds
a) Amino group
b) Carboxyl group
c) A hydrogen atom
d) One of 20 side chains
C. Types (click to go to the amino acid data sheet)
1. There are 20 common amino acids
a) These vary in their side chain
2. Amino acids can be classified according to its side chain characteristics
a) Hydrophobic aliphatic
b) Hydrophilic aliphatic
c) Hydrophobic aromatic
d) Hydrophilic aromatic
e) Sulfur containing
(1) Theses include cysteine and methionine
(2) Cysteine is capable of forming a disulfide bond with and second cysteine
(a) Disulfide bonds are the only common covalent bond between amino acid
side chains
f) Cationic
g) Anionic
3. Essential amino acids
a) Humans can make 11 amino acids from carbohydrates and nitrogen
b) The other amino acids are not made by humans and must be ingested
(1) These are termed the essential amino acids
(2) These are mostly the amino acids with hydrophobic side chains
II.PEPTIDE
BONDS
A. Amino acids in peptides are covalently linked
1. This bond is referred to as a peptide bond
2. It is formed by a dehydration synthesis reaction catalyzed by ribosomes
a) The amino group of one amino acid is linked to the carboxyl group of
another amino acid
B. Hydrolysis of peptide bonds
1. 1 M HCl at high temperatures will hydrolyze peptides into free amino acids
2. Proteases or proteolytic enzymes will enzymatically hydrolyze peptides
III.PRIMARY
STRUCTURE
A. Proteins have four level of organization: primary, secondary, tertiary, and
quaternary
B. The primary level of organization is simply the order of amino acids in the peptide
chain
1. Example: Phe-Ala-Met-Leu-Gln-Trp-Glu-Ile
2. The amino acid sequence is often deduced from the sequence of nucleotides
that code for it
a) It is generally simpler to sequence DNA than it is to sequence proteins
3. The primary structure of protein (order of amino acids) determines how the
protein folds and interacts at the other levels of interaction
a) A protein is about 3.61 Å long per amino acid in an unfolded state
(1) Proteins have no biological activity in an unfolded state
C. Proteins are polymers of amino acids in which the  carbon atoms and peptides
alternate forming the backbone and with specific amino acid side chains projecting
from the  carbon
1. Peptide bond has partial double bond characteristics therefore it is rigid
2. -carbon bonds are flexible
a) Therefore, there is free rotation
D. Polarity
1. Like DNA with its 5' and 3' end, proteins also have polarity
2. Proteins have an amino terminal (N-terminus) end and a carboxyl terminal (Cterminus) end
a) Amino acid sequence is conventionally written from N-terminus to Cterminus
IV.RULES
FOR FOLDING INTO HIGHER LEVELS OF COMPLEXITY
A. The three dimensional folding of a protein is a result between the interactions
described below
1. The peptide bond has partial double bond characteristics and are thus planer
and rigid
2.
3.
4.
5.
6.
V.
a) There is a limit to how much the backbone of the peptide chain is able to
bend
b) The bonds around the -carbon are flexible
Amino acids of the same charge tend to extend the chain and limit its folding
Side chains cannot overlap
Water soluble protein have polar and charged amino acids sticking outward to
interact with water molecules
a) The hydrophobic amino acids are on the inside and do not interact so as not
to interact with water
b) The opposite is true of integral membrane proteins
Hydrogen bonds tend to form between the carbonyl oxygen of one peptide
group and the hydrogen attached to the nitrogen in another peptide group
Sulfhydryl groups tend to form double-covalent bonds
a) This is called a disulfide bond
b) Normally forms between cysteins
SECONDARY STRUCTURE
A. Definition
1. Secondary structures are ordered 2-dimensional structures formed due to
hydrogen binding between peptide groups (hydrogen of amine groups and
oxygen of the carbonyl groups)
a) These hydrogen bonds involve only the peptide backbone, not the side
chains
2. The two common secondary structures are the -helix and the -sheet
B. -helix
1. Each peptide group forms 2 hydrogen bonds
a) The oxygen of the carbonyl group will hydrogen bond to the amine hydrogen
three amino acids in front of it
b) The H of the amine group will hydrogen bond to the oxygen of the carbonyl
group three amino acids behind it
2. There is 3.6 amino acids per turn
a) This is tighter than the DNA double-helix
b) The side chains project outwards from the helix
3. Some amino acids stabilize -helix while others prevent its formation
C. -sheets
1. The peptide chain is almost completely extended but is folded back on itself
2. Hydrogen bonds form between peptide groups of segments lying adjacent and
parallel with one another
a) It is referred to as parallel if both region are going the same direction
b) It is referred to as antiparallel when the regions are going in opposite
directions
3. The amino acid side chains alternate facing above and below the backbone
VI.TERTIARY
STRUCTURE
A. Definition
1. Tertiary structure involves the three-dimensional folding of a protein due to
interactions of amino acid side chains
2. Where secondary structure was a result of hydrogen bonds between peptide
groups, tertiary structure is a result of side chains interactions
a) Interactions
(1) Ionic
(a) Between oppositely charged polar amino acid side chains
(2) Hydrogen bonds
(a) All amino acids except the hydrophobic ones
(3) Hydrophobic interactions
(a) Phenylalanine, leucine, isoleucine, and valine
(4) Disulfide bonds
(a) Cysteine
B. General rules governing interaction of side chains
1. Side chains cannot overlap
a) Some amino acids have bulky side chains that can take up a lot of space
2. Like charges tend to extend the chain
a) Proteins with either large amounts of cationic or anionic side chains will be
longer than proteins carrying a mix of ionic side chains
3. Amino acids with polar side chains are on outside
a) These are hydrophilic and will hydrogen bond with water to keep the protein
soluble
b) This is not true of integral membrane proteins
(1) These will have hydrophobic amino acids on the outside to react with the
hydrophobic fatty acids in the membrane
4. Hydrophobic amino acids tend to cluster together in the center of the protein
a) Hydrophobic amino acid side chains do not interact with water or other
hydrophilic molecules
b) Some peptides have hydrophobic amino acids clustered on the outside, but
these will usually associate with other peptides that also have hydrophobic
patches on the outside
c) This is not true of integral membrane proteins
(1) These will have hydrophilic amino acids clustered together to hide them from
the hydrophobic fatty acids in the membrane
5. Sulfhydryyl groups tend to form double-covalent bonds (disulfide bonds) with
other sulfhydryl group
a) Cysteine is commonly involved in sulfhydryl bonds
b) This constrains structure of the protein
6. 3D shape of a polypeptide chain is a result of a balance between all the
tendencies just described
C. Fibrous proteins
1. -helices and -sheets tend to make the polypeptide rigid
2. Tend to be long and thin
3. Are usually structural proteins
4. Examples: cytoskeleton proteins, elastin, collagen
D. Globular proteins
1. Contain only short -helices and -sheets interspersed with randomly coiled
regions
2. Compact, spherical, and flexible
3. These usually have enzymatic activity
VII.QUATERNARY
STRUCTURE
A. Definition
1. Quaternary structure involves the association of two or more polypeptides into
functional proteins
B. The association often involves hydrophobic interactions
1. Generally hydrophobic amino acids are internal for soluble protein but do
occasionally occur externally
2. Proteins with large hydrophobic patches may reduce its contact with water by
bonding to hydrophobic amino acids of another such protein
a) Self aggregating
C. Definition
1. Number of polypeptides in a protein
a) Dimers, trimers, tetramers, up to 50 or more
2. Types of polypeptides in a protein
a) Homodimers, etc
b) Heterodimers, etc
D. Examples
1. RNA polymerase
a) 5 subunits, four different
2. Hemoglobin
a) 2 alpha and 2 beta subunits
3. DNA polymerase
a) 10 different subunits
VIII.DOMAINS
AND SHAPES
A. Domains
1. Independently folded regions of the protein
a) May be separated by a single break of the polypeptide chain
b) Homology
(1) Many different proteins have similar domains
(a) These often belong to the same gene family
(i) Examples include the Ig-family and DNA binding proteins
B. Shapes
1. Globular
a) Regulatory and enzymatic proteins
b) The binding portions of these proteins are formed from amino acids from
distant parts of the polypeptide chain brought together via tertiary structure
2. Fibrous
a) Structural proteins
IX.ACTIVE
SITES / BINDING SITES
A. Binding site
1. Formed from amino acids from distant parts of the polypeptide brought together
by tertiary and quaternary structures
a) Many interchain bonds bring these amino acids together, therefore, mutation
causing amino acid substitution elsewhere can disrupt protein activity
B. Binding
1. Highly specific interactions between groups of their amino acid side chains and
other molecules
a) Highly specific
(1) May differentiate between sugars (e.g., glucose dehydrogenase binds
glucose, not fructose), nucleic acid sequences (e.g., RNA polymerase binds to
the sequence TAATAT), amino acids (e.g., aminoacyl-tRNA synthetases load
the correct amino acids onto tRNA molecules)
2. The binding usually involves weak bonds, therefore, the molecules must have
complimentary shapes
a) Interactions often involve weak bonds
(1) Opposite charges
(2) Similar hydrophobicity patterns
(3) Hydrogen bonding
b) Substrate with wrong charge, degree of polarity, or shape will not bind
3. The binding site of the protein is complimentary to the object bound
a) Different charges line up, similar hydrophobic patterns, shape, hydrogen
bonding capability
(1) Substrate with wrong charge, shape, or degree of polarity will not fit
C. Regulatory sites
1. Located outside the active site
a) Binds regulatory molecules
2. Binding changes shape of protein, hence changing its activity
X.
DENATURATION
A. Definition
1. Destruction of secondary, tertiary, and / or quaternary structures
2. Primary structure remains intact
a) Without higher levels of organization the protein will have no biological
activity
B. Causes of denaturation
1. Anything that destroys H-bonds, ionic bonds, or hydrophobic interactions will
denature a protein
a) This includes pH, heat, salts, urea
2. Disulfide bonds can be broken with reducing agents such as mercaptoethanol
3. Example of pH disrupting structure
a) Polyglutamic acid (a protein made of just glutamic acids) is a pure alpha
helix at pHs below 5
(1) The carboxyl group on the side chain is not ionized
b) Above pH 6 the alpha helix does not form
(1) The side chain becomes ionized and the electrostatic repulsion destroys the
alpha helix
C. Renaturation
1. May occur spontaneously
2. May not occur at all
a) Reasons why renaturation may not occur
(1) The protein may fold temporally as it is produced (N - C termini)
(2) Molecular chaperones may be needed
(3) Prosthetic group may be lost
D. Hydrolysis
1. 1 M HCl at elevated temperatures
a) This destroys tryptophan, making determination of amino acid composition
difficult
2. Digestive enzymes
a) Exopeptidases
(1) Carboxypeptidases cleave from the C-terminus
(2) Aminopeptidases cleave from the N-terminus
b) Endopeptidases
(1) Cleave at specific side of specific amino acids
XI.REGULATION
OF ENZYMES
A. Regulatory site
1. outside of active site
2. when bound by regulatory molecule, shape of protein changes, hence changing
it’s activity
B. Inhibition
1. End product inhibition
2. A (substrate) binds to active site – B (product) binds to regulatory site
a) B results in conformational change that prevents A from binding