Biochemistry Ch. 7 88-110
Structure-Function relationships in Proteins
Globular Proteins – soluble in aqueous medium and resemble irregular balls
Fibrous Proteins – linear, arranged in a single axis, and repeating unit structure
Transmembrane Proteins – one or more regions cross lipid bilayer
-Primary structure is the linear sequence of amino acids joined by peptide bonds
-Secondary structure is a recurring pattern of regular structure such as α-helix or B-sheet
-Tertiary structure is overall 3D structure of a protein
-Quaternary structure is association of multiple polypeptide subunits by many interactions
Requirements of 3D Structure1. Creation of a binding site specific for just one or few molecules with similar structure
a. Some rigidity in structure is essential for stale structure of binding sites, but still
required to be flexible and mobile
2. Must have an external surface appropriate for its environment (polar, non-polar residues)
The 3D Structure of Peptide Backbone – amino acids joined by peptide bonds of carboxyl group and
amino group of next amino acid
-peptide bond assumes a trans configuration in which successive α-carbons and R groups are located on
opposite sides of peptide bond
-peptide bond has two resonance structures and therefore the backbone consists of rigid planes formed
by peptide groups, but still allowing rotation around torsion angles that occur around the bond between
α carbon and α amino group and around bond between α carbon and carbonyl group
Secondary Structure – α-helix and B-sheet are secondary structures formed by repeating elements
1. The α-helix – popular in globular proteins, membrane spanning domains, and DNA-binding
domains that maximizes H-bonding while maintaining rotation angles
a. Formed by H-bonds between carbonyl oxygen and amide hydrogen 4 residues down
b. Trans- side chains of amino acids project backward and outward from helix to avoid
steric hindrances
c. Proline CANNOT form within an α-helix
2. B-Sheets – maximize H-bonding between backbones while maintaining torsion angles
a. H-bonding occurs between regions of separate neighboring peptide strands aligned
parallel to one another
b. Carbonyl oxygen bonded to amide hydrogen on adjacent strand
c. Optimal binding occurs when the sheet is bent
d. Parallel B-sheets are when peptide strands run in same direction
e. Antiparallel B-sheets are when peptide strands run in opposite directions
f. Side chains alternate going above and below the sheet
3. B-turns – non-regular structure involving short regions that are usually 4 successive amino acids
and involve abrupt turn and connect antiparallel B-sheets
a. –surface of large globular proteins have at least one omega loop
Patterns of Secondary Structure – Lactate dehydrogenase is typical globular protein with α-helices and
B-sheets that are mostly similar in nature (α-helix 12 residues, B-sheets 6 residues)
-motifs – relatively small arrangements of secondary structure that are recognized in many proteins,
such as repeating patterns of B α B α B structural motif
-some connecting residues that connect different α-helices or B-sheets are recognized like omega loop,
but irregular regions are called coils – not random, but stabilized through H-bonds and do not vary from
one protein molecule to another
-coils, loops, and other segments are usually more flexible than α-helices or sheets
Tertiary Structure – folding pattern of the secondary structural elements into 3D conformation which
serves all aspects of a protein’s function
-creates specific and flexible binding sites for ligands, maintains residues on surface appropriate for
protein’s location (polar residues for cytosolic proteins and hydrophobic for intramembranous)
-forces that maintain tertiary structure are van der Waals interactions, H-bonds, ionic bonds,
hydrophobic effect, and disulfide bond formation
Domains in Tertiary Structure – tertiary structure can be described by regions called structural domains
-each domain formed from continuous sequence of AA that are folded into 3D space independently.
Folds in Globular Proteins – large patterns of 3D structure that have been organized in proteins
associated with a characteristic activity (ATP binding and hydrolysis, or NAD_ binding)
1. The Actin Fold – four subunits of actin contribute to folding pattern known as actin fold. ATP is
bound to the middle in the fold by AA domains on both sides, and when ATP binds a
conformational change closes the middle cleft and ATP  ADP
a. Found in actin, heat-shock protein 70, hexokinase
2. Nucleotide Binding Fold – fold can be formed through one domain, such as the one from
Lactate Dehydrogenase: domain 1 forms a nucleotide binding fold, a binding site for NAD+, or in
other proteins for molecules of similar structure like riboflavin
a. NAD+ and NADP+ have drastically different nucleotide binding folds
Solubility of Globular Proteins in Aqueous Environment – most globular proteins are soluble in the cell
and contain a core of hydrophobic amino acids (Val, Leu, Ile, Met, Phe) that is densely packed to
maximize van der Waals forces
-charged, polar AA are found on the surface of the protein where they form ionic pairs with solvent and
often bind inorganic ions (K+, PO4, Cl-)
-charged amino acids on the inside usually involved in forming specific binding sites
Tertiary Structure of Transmembrane Proteins – transmembrane proteins such as B2-adrenergic
receptor contain membrane spanning domains and intra-/extra- cellular domains
-ion channels, transport proteins, neurotransmitter receptors, and hormone receptors contain
membrane-spanning segments that are α-helices with hydrophobic residues exposed to lipid bilayer and
connected by loops with hydrophilic AA side chains that extend into aqueous medium on both sides of
membrane
- B2-adrenergic receptor has helices clump together to form binding site for epinephrine
-transmembrane receptors are post-translationally modified such as addition of branched highmannose oligosaccharides or anchoring by palmitoyl groups, or phosphorylation sites, etc…
Quaternary Structure – association of individual polypeptide subunits in a specific manner, such as
dimers, tetramers, oligomers to make one functional protein
-homo, heterodimer, trimer, etc… used to describe structure, also multimer, oligomer, etc…
-protomer – unit structure within protein composed of multiple non-identical subunits
-adult Hgb is A2B2, and one AB pair is considered protomer
-contact region between subunits resemble interior of a single subunit: closely packed hydrophobic AA,
H+bonds involving polypeptide backbones and side chains, and occasional ionic bonds
-subunits of globular proteins rarely held together by disulfide bonds and never by other covalents
-subunits of fibrous proteins are frequently linked together through covalent bonds
-A multisubunit structure increases the stability of the protein and the increase in size increases
interaction between AA residues to make it more difficult to unfold and refold
-multisubunit structure will allow for cooperative binding between subunits in binding ligands or to form
binding sites with high affinity for large molecules
Insulin – composed of 2 non-identical chains attached to each other through disulfide bonds.
-it is synthesized as a single chain forms a disulfide bond with itself and is later cleaved into 2 subunits
Quantitation of Ligand Binding – folding of a protein can create binding site for ligand such as NAD+ or
ATP and adrenaline: binding affinity measured using association constant Ka
** L + P  LP (K1 is constant in forward direction, K2 is constant in reverse direction)
** Keq = k1/k2 = [LP] / [L][P] = Ka = 1/Kd
Kd = dissociation constant, L = ligand, P = protein, Ka = association constant, Keq = equilibrium constant
Ka = k1 for association of ligand with binding site divided by rate constant k2 for dissociation of LP
-tighter the binding of ligand to protein, the higher is the ka and lower the kd
Structure-Function Relationships in Hemoglobin/Myoglobin – both are O2 binding proteins with similar
protein structures, except myglobin has 1 chain and hemoglobin is a tetramer of alpha and beta subunits
(AB protomer), which each subunit having homology to myoglobin
-hemoglobin’s tetrameric structure saturates with O2 in lungs and releases in tissue
-curves showing O2 saturation against PO2 suggest myglobin has a hyperbolic curve and hemoglobin has
a sigmoidal curve
-when PO2 is high in lungs, both myoglobin and hemoglobin saturated with oxygen
-when PO2 is low in tissues, hemoglobin cannot bind O2 as well as myoglobin
-myoglobin in heart and skeletal muscle can bind O2 released by hemoglobin for ATP 
myglobin releases O2 which is picked up by cytochrome oxidase for ATP generation
O2 Binding and Heme – tertiary structure of myoglobin is 8 alpha helices with short coils  globin fold
and has no B-sheets  helices create hydrophobic O2 binding pocket with heme and Fe2+
Heme – porphyrin ring composed of 4 pyrrole rings linked by methylene bridges binding an Fe2+ atom
-organic, tightly bound ligands like heme of myoglobin are called prosthetic groups, and a protein with
its attached prosthetic group is called a holoprotein  without which it is called apoprotein
Fe2+ can bind 6 ligands, 2 of which stabilize its binding to porphyrin ring and 4 for O2 or CO2 binding
-Myoglobin is released from skeletal/cardiac muscle upon cell damage and passes into the blood to be
excreted in the urine, and can turn urine red (myoglobinuria)
Cooperativity of O2 binding in Hemoglobin – coopaterativity of O2 binding to hemoglobin caused by
conformational changes in tertiary structure that takes place when O2 binds from the T (tense) state
with low O2 affinity to the R (relaxed) state with a high affinity for O2
-binding affinity for first O2 is really low, but when 1 binds, the next one will bind much easier
-Sickle Cell Anemia – caused by wrong quaternary structure where polymerization of sickle cell
hemoglobin (Hbs) into long fibers distort shape of RBC due to SNP in the B-chain of hemoglobin, and
polymerizes until long fibers are formed
Structure-Function Relationships in Immunoglobulins – antibodies act as defense against foreign
antigens by binding to them and initiating the destruction process
-all of them have similar structure with 2 small light chains and two identical heavy chains joined by
disulfide bonds
-there are 5 major classes of immunoglobulins, most abundant of which is γ-globulin (IgG class) and both
its light and heavy chains consist of domains known as immunoglobulin fold: collapsed B-sheets known
as a B-barrel
-both heavy and light chains contain variable [C] regions which interact to produce a single antigenbinding site at the branches of the Y-shaped molecule
-each population of B cells forms an antibody with a different AA composition in the variable region and
the dissociation constant is very small (VERY TIGHT BINDING)
-the constant (Fc) regions are important for antibody-antigen complex binding to phagocytic cells
Amyloidosis – amyloid is formed from degradation products of lambda or kappa-light chains that
deposit most frequently in extracellular matrix of kidney and heart and sometimes tongue
-in other amyloidosis, proteins arise in characteristic organ such as in rheumatoid arthritis,
where it is derived from serum protein called serum amyloid A produced in the liver in response
to inflammation and deposits in the kidney
Protein Folding – although peptide bonds are rigid, flexibility in backbone allows many conformations,
but proteins generally assume the same conformation called the native conformation
**Primary structure determines folding into a protein’s 3D conformation, meaning that sequence of AA
side chains dictate folding into tertiary and quaternary structure
-denatured proteins can sometimes refold into their native conformation
-in the cell, not all proteins can refold on their own into their native conformation, and first go into
many high-energy conformations to slow the process kinetic barriers
-heat shock proteins use energy provided from ATP hydrolysis to assist in folding
-hsp70 proteins bind to nascent polypeptide chains to keep them from being folded
prematurely, and also unfold proteins before entering membrane of mitochondria
-hsp60 is a chaperonin – or barrel shaped protein where unfolded protein fits into cavity and
serves as template for folding process
-cis-trans isomerases – converts trans peptide bond preceding proline into cis conformation for hairpin
turns
-protein disulfide isomerase participate in folding by breaking and reforming disulfide bonds between
two cysteine residues in transient structures during folding
Protein Denaturation
1. Denaturation through nonenzymatic modification of proteins – AA can be chemically
modified not through enzymes like glycosylation or oxidation
a. Glycosylation involves glucose n blood binding to exposed AA on protein to form
irreversible glycosylated protein
i. Proteins that turn over slowly like collagen and hemoglobin are glycosylated
ii. Rate of glycosylation only proportional to amount of glucose in blood,
therefore hyperglycemic people have more glycosylated proteins
b. Oxidation of collagen and other proteins form aggregates and cross-links referred to
as advanced glycosylation end products (AGEs)
i. AGEs accumulate with age even in individuals with normal blood glucose
2. Denaturation by Temp, pH, and Solvent – all of these disrupt ionic, H- bonds and
hydrophobic bonds
a. At low pH, ionic bonds and H-bonds would be disrupted, and at high pH, H-bonds on
basic amino acids would be disrupted
b. Proteins denatured by gastric juice (pH 1-2), but it cannot break peptide bonds, only
make it a better substrate for digestive enzymes
c. Temperature increases vibrational and rotational energies of bonds to destabilize
tertiary structure
d. Proteins can also be denatured by amphipathic agents such as urea, guanidine
hydrochloride, or SDS
e. Hydrophobic molecules can also denature proteins by disturbing hydrophobic
interactions in the protein
3. Protein Misfolding and Prions – prions are believed to cause neurodegenerative disease by
acting as a template to misfold other cellular prion proteins into a form that cannot be
degraded
a. Prion – proteinacious infectious agent and can be acquired through infection (Mad
cow disease) or from inherited mutations (Creutzfeldt-Jakob disease [CJD])
i. Attributed to cannibalism
ii. Prion diseases are classified as transmissible spongiform encephalopathies
characterized by spongiform degeneration and astrocytic gliosis in CNS,
plaques are normally seen that are resistant to degradation
-prion on brain is encoded by normal human genome, but the prion is folded differently that
allows it to aggregate.
-Normal protein is called PrPc and disease is PrPsc, but PrPsc has enriched B-sheet
structure compared to PrPc (40% alpha helix)
-disease occurs when PrPsc dimers are ingested into the body that act to lower
activation energy for the normal conformers and cause them to also assume the prion
state
Familial Prion Diseases are caused by SNPs in gene encoding the Pr protein, and presents in diseases
like Creutzfeldt-Jakob Disease, which is autosomal dominant and presents in fourth decade of life
-sporadic form accounts for 85% while inherited, familial form accounts for 15%
Myocardial Infarction – specific heart proteins such as creatine kinase and cardiac troponin are
measured in the blood for evidence of cardiac muscle damage