Protein Structure

Part 1 Overview

• Introduction to protein structure

• Energetics of protein folding and effects of water

• Peptide bond characteristics

• Secondary structures:

  • Alpha helices

  • Beta sheets

  • Beta turns

Fundamentals of Protein Structure

Importance of Structure

• The function of a protein is directly dependent on its structure.

• The three-dimensional (3D) structure of a protein is dictated by its amino acid sequence.

• A native protein typically has a limited number of stable conformations (often 1, 2, or 3 structures).

• Key forces that determine protein structure are predominantly non-covalent and weak interactions.

• Despite the vast diversity of protein structures, there are common structural patterns.

Useful Definitions

• Conformation: The spatial arrangement of atoms in a structure resulting from rotation about C-C single bonds.

• Native Protein: Proteins in their functional, folded state.

• Protein Stability: The tendency of a protein to maintain its folded state.

• Folding: The process by which a protein acquires its three-dimensional configuration.

Stabilizing Interactions in Proteins

Overview of Interactions

• Protein stability arises from numerous weak chemical interactions.

• The backbone (primary structure) of a protein is formed by covalent bonds, notably peptide bonds.

• The 3D structure of proteins hinges on a variety of bond types, including:

  • Van der Waals interactions

  • Hydrogen bonds

  • Ionic bonds (also known as "salt bridges")

  • Hydrophobic interactions

  • Disulfide bonds

Energetics of Protein Folding

Reaction Direction

• Factors determining the direction of a reaction:

  1. $ΔG^o = -RT ext{ln} K_{eq}$

  2. $ΔG = ΔH - TΔS$

• Reactions proceed in the direction where $ΔG$ is negative. This requires:

  • $ΔH$ to be negative (indicating a lower energy state)

  • $ΔS$ to be positive (indicating increased entropy or randomness)

  • Note: $T$ is the absolute temperature in Kelvin, always positive.

Role of Water in Protein Folding

• Water significantly influences protein folding.

• Presence of a hydrophobic group in an aqueous environment causes it to immobilize an ice-like "shell" of water molecules (termed clathrate).

• This immobilization decreases entropy due to reduced randomness in the system.

• If hydrophobic groups cluster, their surface area diminishes, leading to fewer water molecules in the organized state.

• Consequently, water molecules drive hydrophobic side chains into the protein's center, facilitating protein folding.

Energetics Equation

• The folding equation is presented as $ΔG = ΔH - TΔS$.

• A negative $ΔG$ is vital for folding to happen:

  • Folding reduces the entropy of the protein itself, which could be unfavorable.

  • Enthalpic contributions arise as hydrogen bonds and ionic interactions lower the overall energy state (favorable).

  • The entropy of the surrounding water increases when the protein folds, further contributing to a favorable outcome.

  • Hence, the net $ΔG$ becomes negative, promoting protein folding as a favorable overall process.

Protein Folding Conceptual Model

• Protein folding can be conceptualized as an "energy funnel", with directions of folding leading to lower energy states.

Complexity of Protein Folding

• Protein folding is an intricate process influenced by various factors and hence cannot be overly simplified.

Molecular Chaperones in Protein Folding

Role of Chaperones

• Molecular chaperones are specialized proteins that assist in the proper folding of other proteins.

Rules Governing Protein Folding

Key Rules

1. Hydrophobic residues are generally hidden within the protein's interior, increasing $ΔS$ (entropy).

2. Maximization of weak interactions within the protein chain typically occurs, which reduces $ΔH$ (enthalpy).

Misfolded Proteins and Prions

Introduction to Prions

• Prions represent unique infectious agents responsible for various diseases, such as “mad cow disease.”

• They consist of misfolded proteins that induce misfolding in neighboring proteins, therefore acting infectively.

Levels of Protein Structure

Primary Structure

• The primary structure of a protein is defined as the linear sequence of amino acids linked by peptide bonds, with distinct ends:

  • Free amino group (N-terminus)

  • Free carboxyl group (C-terminus)

Peptide Bond Characteristics

Properties

• The peptide bond displays resonance between two extreme structures, giving it partial double bond characteristics, which result in rigidity and planarity.

• The carbonyl oxygen carries a partial negative charge; the amide nitrogen possesses a partial positive charge, forming a dipole moment.

• The rigidity implies that rotation around peptide bonds is restricted.

Configuration

• Almost all peptide bonds are found in the trans configuration.

Allowed Rotations

Peptide Bond Rotation

• Rotation is permitted around the N-Cα and Cα-C bonds only.

• The N-Cα bond angle is referred to as $ϕ$ (phi), while the Cα-C bond angle is termed $ψ$ (psi).

Steric Hindrance Constraints

• Steric hindrance, particularly between adjacent carbonyl oxygens, limits feasible $ϕ$ and $ψ$ angles.

Ramachandran Plots

• Ramachandran plots illustrate the allowed and forbidden angles for $ϕ$ and $ψ$, with white areas indicating impractical configurations.

Protein Secondary Structure

Overview

• The secondary structure relates to particularly stable local conformations formed by adjacent amino acids within a polypeptide.

Alpha Helices

• The alpha helix is a common secondary structure whose existence was initially theorized and later confirmed through X-ray diffraction.

• The peptide backbone coils around an imaginary axis with a right-handed twist, accommodating approximately 3.6 amino acids per turn.

• Hydrogen bonds stabilize the alpha helix between the ith and (i+4)th amino acids, aligning parallel to the helix axis.

Destabilizing Factors

• Certain amino acids can disrupt the stability of alpha helices:

  • Proline disturbs it due to its unique side chain connection to the amino group.

  • Glycine is less frequent in alpha helices due to its flexible conformation.

  • Sequential runs of positively (Lys, Arg, His) or negatively (Glu, Asp) charged residues can destabilize the helix due to repulsive interactions.

  • Bulky side chains like those of Thr, Leu, Asn, Ser, and Cys may also cause destabilization if too close due to steric hindrance.

Beta Structure

• The beta conformation organizes peptide chains into beta sheets, which were also first theorized and later substantiated by X-ray crystallography.

• Peptide strands are fully extended and arrayed in sheets, characterized by two arrangements forming pleated structures.

• Small R groups alternate and extend from opposite faces of the beta sheet.

Antiparallel Beta Sheets

• In antiparallel beta sheets, individual beta strands maintain a fully extended zig-zag conformation.

• Strands run adjacent and in opposite directions, held together by hydrogen bonds, with a repeat distance of approximately 7 Å.

• In this arrangement, the hydrogen bond donor and acceptor atoms align in a straight line, strengthening the bonds.

Parallel Beta Sheets

• For parallel beta sheets, strands remain extended but share the same direction.

• Hydrogen bonds persist but with a repeat distance around 6.5 Å, and the donor and acceptor hydrogen bonding atoms do not align straightforwardly, resulting in weaker bonds.

Other Secondary Structures

• Beta turns, linking the ends of antiparallel beta sheets, and collagen, with a distinct triple-helical structure due to a recurring amino acid sequence of Gly-X-Y (X and Y often being proline or hydroxyproline), are notable secondary structures.

Amino Acids and Secondary Structures

• Different amino acids exhibit varying tendencies to participate in specific secondary structures, such as alpha helices, beta conformations, and beta turns.

• Common examples include:

  • Glutamic acid (Glu) and Methionine (Met) favoring alpha helices, while others are more suited to beta conformations or beta turns, e.g., Glycine (Gly) and Proline (Pro).

Review Points

• Protein folding is primarily influenced by non-covalent interactions.

• Both enthalpic (favorable) and entropic (unfavorable) contributions are integral to the folding process.

• The rigidity and planarity of the peptide bond are due to its structural properties, resulting in restricted rotations and configurations which are graphically represented in Ramachandran plots.

• Alpha helices and beta sheets represent common and essential secondary structures in proteins, with varying amino acid compositions impacting their stability and formation.