41 - Protein Folding

Primary Structure of Proteins

The linear sequence of amino acids in a polypeptide chain linked by peptide bonds

Importance of Primary Structure

The primary sequence provides the blueprint that determines how a protein folds into secondary, tertiary, and quaternary structures

Side Chains (R-groups) Role in Folding

Amino acid side chains have diverse chemical properties (hydrophobic, polar, charged) that drive folding through intermolecular interactions

Secondary Structure

The local spatial arrangement of main-chain atoms in a polypeptide segment without considering side-chain positioning

Tertiary Structure

The overall 3D folding of a single polypeptide chain primarily driven by side-chain interactions

Quaternary Structure

The arrangement and interaction of multiple polypeptide chains within a functional protein complex

Resonance Stabilization of Peptide Bond

Electrons in the peptide bond are delocalized, giving it partial double-bond character and restricting rotation

Amide Plane

The rigid planar structure formed around the peptide bond due to resonance stabilization

Peptide Bond Rigidity

The peptide bond cannot freely rotate due to its partial double-bond character

Omega (ω) Angle

The dihedral angle describing rotation around the peptide bond; typically fixed at approximately 180° (trans configuration)

Trans Peptide Bond Configuration

The peptide bond most commonly exists in the trans configuration (ω ≈ 180°), minimizing steric hindrance

Phi (φ) Angle

The dihedral angle representing rotation between the nitrogen atom and the alpha carbon (N–Cα bond)

Psi (ψ) Angle

The dihedral angle representing rotation between the alpha carbon and carbonyl carbon (Cα–C bond)

Rotational Freedom in Backbone

Only the φ and ψ bonds rotate freely; peptide bonds remain rigid

Steric Hindrance in Protein Folding

Physical crowding between atoms restricts possible φ and ψ angle combinations

Ramachandran Plot

A graphical representation showing sterically allowed combinations of φ and ψ angles in protein structures

Purpose of Ramachandran Plot

Used to predict and visualize allowed backbone conformations based on steric constraints

Allowed Conformations in Proteins

Only specific φ and ψ angle combinations are possible due to steric hindrance and tetrahedral geometry of carbon

Random Coil

A segment of polypeptide lacking a regular repeating secondary structure pattern

Regular Secondary Structure

Regions where φ and ψ angles remain relatively constant throughout the structure

Alpha Helix

A right-handed helical secondary structure stabilized by internal hydrogen bonds along the backbone

Hydrogen Bonds in Alpha Helix

Stabilizing bonds occur between the carbonyl oxygen of residue i and the amide hydrogen of residue i+4

Residues per Turn in Alpha Helix

One complete turn of an alpha helix contains approximately 3.6 amino acid residues

Orientation of R-Groups in Alpha Helix

Side chains project outward from the helix backbone

Helical Wheel Projection

• A 2D representation of an alpha helix used to visualize spatial arrangement of amino acid side chains

• good for determining properties of it

Stabilization of Alpha Helix

Stabilized by optimal internal hydrogen bonding and favorable side-chain interactions

Alpha Helix Dipole Moment

The alignment of peptide bond dipoles creates partial positive charge at the N-terminus and partial negative charge at the C-terminus

Directionality of Alpha Helix

Amide NH groups point toward the N-terminus and carbonyl groups point toward the C-terminus

Amino Acid Propensity for Alpha Helix Formation

Certain amino acids have higher tendencies to form alpha helices based on energetics

Alanine Helix Propensity

Alanine has the highest tendency to form alpha helices and is used as a reference point

Proline as Helix Breaker

Proline disrupts alpha helices due to its rigid cyclic structure that restricts backbone flexibility

Glycine in Alpha Helices

Glycine destabilizes alpha helices due to excessive conformational flexibility

Charge Repulsion in Alpha Helices

Adjacent similarly charged side chains (e.g., glutamate residues) repel and destabilize helices

Beta Strand

An extended polypeptide chain forming part of a beta sheet

Beta Sheet

A secondary structure formed when multiple beta strands align side by side

Backbone Structure in Beta Sheets

Polypeptide chains adopt a zigzag conformation

Hydrogen Bonds in Beta Sheets

Form between carbonyl oxygen of one strand and amide hydrogen of an adjacent strand

Parallel Beta Sheets

Adjacent strands run in the same N-to-C direction

Antiparallel Beta Sheets

Adjacent strands run in opposite N-to-C directions

Stability of Beta Sheets

Antiparallel beta sheets are generally more stable and more common than parallel sheets

Tertiary Structure Environment

Protein folding occurs in crowded, aqueous cellular environments containing ions and macromolecules

Conformational Entropy of Unfolded Protein

The unfolded protein has many possible conformations, resulting in high entropy

Entropy Change During Folding

Folding reduces protein entropy, making folding thermodynamically unfavorable on its own

Hydrophobic Effect

The tendency of hydrophobic residues to cluster together, releasing ordered water molecules and increasing system entropy

Driving Force of Hydrophobic Effect

Release of structured water molecules increases entropy of surrounding solvent

Cooperativity in Folding

Initial hydrophobic interactions promote additional folding interactions, creating a zipper-like effect

Polar Backbone Constraints

Backbone polar groups must form hydrogen bonds or structured conformations when buried

Membrane Protein Hydrophobic Regions

Hydrophobic residues interact with lipid bilayers in membrane proteins

Motif (Protein Structure)

A recognizable folding pattern involving two or more secondary structure elements and their connections

Motif Function Prediction

Similar motifs often correspond to similar protein functions

Protein Structural Databases

Collections of protein structures used to relate structure to function

Protein Data Bank (PDB)

A database containing experimentally determined protein structures

Structural Classification of Proteins (SCOP2)

A database categorizing proteins based on structural and evolutionary relationships

Domain (Protein Structure)

A segment of a protein that is independently stable and can function as a single unit

Independent Domain Function

Some domains retain function even after separation from the full protein

Example of Protein with Domains

Troponin C contains separate calcium-binding domains

fibrous proteins

• play structural roles

• are elongated/filamentous in shape

• ex. collagen and keratin

• they’re built to last, not be modified/regulated

• some are regulated: actin and tubulin

  • they form fibres, but aren’t fibrous themselves

globular proteins

• carryout chemical work like synthesis, transport and metabolism

• they’re folded into compact structures

• they bury their hydrophobic aa in the core, and hydrophilic in the surface in contact with (aq) env

• ex. myoglobin and hemoglobin

membrane proteins

• they have a higher proportion of hydrophobic aa

• they use this to interact with the lipid bilayer

• peripheral: sits on the top of the bilayer

• integral: fully integrated into the membrane

• ex. ATP synthase and insulin receptor

forces governing protein folding (5)

1. electrostatic

2. H bonds

3. van der waals

4. hydrophobic effect

5. cooperativity

electrostatic forces

• strong interactions within a vacuum

  • weakened due to water and it’s large dielectric constant

• charge-charge

• long distance

• many atoms in proteins have partial charges, that add up to have a significant impact

hydrogen bonds

• occur between peptide groups and water on a protein surface

• in the core, they’re generated between peptide groups

• H-bonds are bidirectional

van der waals forces

• very short range, significant when molecules are almost in contact

• influential in high numbers

• have corresponding repulsive forces that are strong when atoms are too close

hydrophobic effect

• hydrophobic groups cluster within the core

• they reduce the # of free H-bonds water molecules nearby can make

  • makes it thermodynamically unfavourable

• water becomes too ordered around to hydrophobic molecule and decreases entropy

  • the increase in enthalpy (maintaining H-bond #) results in decrease of entropy

cooperativity

• proteins are cooperative in folding regarding secondary structural elements

• other factors also add on, like the hydrophobic effect

• a sharp transition of a protein is a strong indicator of high cooperation

Collagen Triple Helix Structure

Collagen consists of three polypeptide chains wound into a tight triple helix that gives it high tensile strength

Repeating Amino Acid Pattern in Collagen

Every third amino acid in collagen is glycine, allowing tight packing in the center of the triple helix

Role of Proline and Hydroxyproline in Collagen

Proline and hydroxyproline stabilize the triple helix and help maintain collagen's rigid structure

Vitamin C Role in Collagen Stability

Vitamin C is required to convert proline into hydroxyproline, which is essential for stabilizing collagen

Scurvy and Collagen

Vitamin C deficiency prevents proper collagen formation, leading to weak connective tissues and symptoms such as bruising and tooth loss

Denaturation of Collagen and Gelatin Formation

Heating collagen breaks the triple helix, and upon cooling the denatured chains absorb water to form gelatin

Type I vs Type IV Collagen (Conceptual Difference)

Type I collagen forms strong rope-like fibers, while Type IV collagen forms sheet-like networks in basement membranes

Structure–Function Relationship in Collagen

The repeating glycine–proline–hydroxyproline sequence allows collagen to form strong structural fibers

Importance of Glycine Size in Collagen

Glycine must occupy every third position because its small size allows tight packing within the triple helix, and larger amino acids disrupt structure

Myoglobin (historical significance)

Myoglobin was the first protein whose 3D atomic structure was determined, providing the foundation for understanding protein structure

Primary Function of Myoglobin

Myoglobin stores oxygen in muscle cells for use during periods of high energy demand

Heme Group Function in Myoglobin

Myoglobin contains a heme group with an iron ion that binds oxygen molecules

Location of Heme in Myoglobin

The heme group is buried in a deep pocket within the protein, protecting and stabilizing oxygen binding

Protein Dynamics and Oxygen Binding

Myoglobin is flexible and constantly moving, allowing temporary openings that let oxygen enter and leave the binding site

Hydrophobic vs Charged Residues in Globular Proteins

Hydrophobic amino acids are typically buried inside proteins, while charged amino acids are usually found on the surface

Salt Bridges in Protein Stability

Oppositely charged amino acids on the protein surface can form salt bridges that help stabilize protein structure

Myoglobin Adaptation in Diving Animals

Marine mammals have higher concentrations of myoglobin to store more oxygen for long underwater dives

Surface Charge Adaptation in Whale Myoglobin

Whale myoglobin has more positively charged amino acids on its surface, which helps prevent protein aggregation at high concentrations

Structure–Function Relationship in Myoglobin

The folded globular structure of myoglobin positions the heme group and amino acids in a way that enables efficient oxygen storage

Hemoglobin Structure

Hemoglobin is composed of four protein chains (two alpha and two beta), each containing a heme group that binds oxygen

Function of Hemoglobin

Hemoglobin transports oxygen through the blood by reversibly binding oxygen to iron atoms in its heme groups

Cooperative Binding in Hemoglobin

Binding of oxygen to one subunit causes structural changes that make the remaining subunits bind oxygen more easily

Allosteric Shape Change in Hemoglobin

Oxygen binding causes hemoglobin to change shape, and this structural shift is transmitted between subunits

Hemoglobin Oxygen Loading and Release

Hemoglobin binds oxygen efficiently in the lungs where oxygen is abundant and releases it in tissues where oxygen levels are low

Carbon Monoxide Toxicity and Hemoglobin

Carbon monoxide binds strongly to hemoglobin heme groups, blocking oxygen binding and preventing oxygen transport

Nitric Oxide Transport by Hemoglobin

Hemoglobin can bind and transport nitric oxide, which helps regulate blood vessel relaxation and blood pressure

Sickle Cell Mutation

Sickle cell disease results from a mutation where glutamate is replaced by valine in the beta chain, causing hemoglobin molecules to stick together

Effect of Sickle Cell Hemoglobin on Red Blood Cells

Mutated hemoglobin forms fibers that distort red blood cells into a sickle shape, making them fragile and prone to rupture

Cooperativity Advantage in Hemoglobin Function

Cooperative oxygen binding allows hemoglobin to load oxygen efficiently in the lungs and release large amounts in tissues

Primary Function of ATP Synthase

ATP synthase produces ATP using energy from a proton gradient across a membrane

ATP Synthase as a Molecular Machine

ATP synthase functions as both a molecular motor and an enzyme that converts mechanical energy into chemical energy

F0 Portion of ATP Synthase

The F0 component is embedded in the membrane and rotates when hydrogen ions flow through it

F1 Portion of ATP Synthase

The F1 component generates ATP by undergoing shape changes that bind ADP and phosphate, form ATP, and release it

Energy Source for ATP Production

The flow of hydrogen ions down their concentration gradient provides the energy that drives ATP synthesis

Rotation-Driven ATP Formation

Rotation of the central axle forces conformational changes in the F1 motor that allow ATP synthesis to occur