Lecture 1/13/26 Proteins and Protein Folding

Introduction to Proteins

• Proteins are fundamental biological building blocks of the cell, also referred to as polypeptides.

• The terms "protein" and "polypeptide" are used interchangeably and signify similar concepts.

Structure of Amino Acids

• Proteins are composed of individual units called amino acids.

• Each amino acid has a central carbon atom (alpha carbon) linked to three groups:

  • An amino group (in red)

  • A carboxyl group (in blue)

  • A variable R group (in green) which determines the identity of the amino acid.

• There are 20 different amino acids, resulting in 20 different side chains, contributing to the diversity of proteins.

Formation of Peptide Bonds

• Amino acids link to form proteins via peptide bonds, created through a dehydration synthesis reaction.

• The carboxyl group of one amino acid reacts with the amino group of another, releasing water.

• This process connects two amino acids into a dipeptide, and can repeat hundreds or thousands of times, resulting in larger proteins.

Ribosomes as Protein Factories

• Cellular ribosomes catalyze the peptide bond formation, greatly increasing the speed of protein synthesis.

Primary Structure of Proteins

• The primary structure of a protein refers to its amino acid sequence.

• Example: A sequence with 434 amino acids demonstrates an average protein size found in cells.

Rigidity of Peptide Bonds

• Peptide bonds exhibit rigidity due to resonance involving electrons from oxygen to nitrogen, preventing rotation despite being single bonds.

Clicker Question: Peptide Bonds in Pentapeptide

• To form a pentapeptide (five amino acids), four peptide bonds are required due to each bond linking two amino acids.

Clicker Question: Total Combinations of Amino Acids

• With 20 amino acids, the number of possible combinations for three-position spots is $20^3 = 8000$, illustrating the vast potential for protein diversity.

Concept of Protein Folding

• The primary structure alone is insufficient for protein functionality; proteins require proper folding to become active.

• The relationship structure = function is critical. Without correct folding, proteins may be non-functional.

Stages of Protein Folding

1. Unfolded State: Newly synthesized proteins begin as a linear amino acid sequence without shape.

2. Secondary Structure Formation: Local interactions among nearby amino acids lead to folded regions (alpha helices and beta sheets).

3. Tertiary Structure: The overall 3-dimensional structure is achieved primarily through hydrophobic interactions, where hydrophobic residues are buried in the protein's core to avoid contact with water.

4. Biological Activity: A protein is only biologically active in its tertiary structure; secondary structure alone does not suffice.

Importance of Folding

• Folding is a determining factor in protein activity. Misfolded proteins can become inactive or dysfunctional.

• Chaperone proteins may assist unfolding or the folding process.

Experimental Evidence of Folding Importance - Anfinsen's Experiment

• Dr. Anfinsen's research utilized the small protein ribonuclease, studying the link between its amino acid sequence and its active conformation.

• Active Ribonuclease: Initially, the active protein was fully folded with active sites intact. Upon treatment with a reducing agent and urea, disulfide and non-covalent bonds were disrupted, yielding an inactive protein.

• Refolding Experiment: Removing the denaturing agents allowed the protein to refold and regain 100% activity, demonstrating that primary structure determines final folding.

Chemical Bonds Stabilizing Tertiary Structure

• The overarching bonds supporting tertiary structure are non-covalent, including:

  • Van der Waals interactions

  • Hydrogen bonds

  • Ionic bonds

  • Hydrophobic effects

• These bonds are collectively weak but powerful due to their large numbers, resulting in significant stabilization of the protein structure.

Conformational Changes and Protein Activity

• Conformational changes are reversible and crucial for protein functionality; they can switch activity states (on/off).

• Example: Activation of proteasome complex requires a conformational change that separates subunits.

• Binding of substrates or ligand triggers these conformational changes, rather than random events.

Diverse Interactions of Amino Acid Types

• Amino acids can be classified into various groups, relevant to their charge and hydrophilicity:

  1. Polar Charged (Hydrophilic): Form ionic bonds (e.g., aspartic acid, glutamic acid, lysine).

  2. Polar Uncharged: Capable of forming hydrogen bonds, often involved in protein modifications like phosphorylation (serine, threonine, tyrosine).

  3. Nonpolar (Hydrophobic): Typically found in protein cores, away from water environments.

• Glycine, cysteine, and proline do not fit standard classifications and have unique roles in protein structure.

Phosphorylation: A Key Modification

• Phosphorylation, often mediated by kinases, adds a phosphate group to the hydroxyl in serine, threonine, and tyrosine.

• This modification can change protein folding and activity, often resulting in activation.