Protein Secondary Structure Comprehensive Study Guide

Course Overview and Learning Objectives

• Goal of the Presentation: To explore the protein secondary structure within the context of CHEM 375.

• Specific Learning Objectives:
      * Develop an understanding of how the inherent chemical characteristics of amino acids and the nature of peptide bonds facilitate the formation of specific secondary structures.
      * Analyze the Ramachandran plot to understand its relation to secondary structure and determine why only a limited set of $\Phi$ and $\Psi$ angles are energetically allowed.
      * Describe the specific hydrogen bonding patterns and structural features defining the two primary secondary structures in proteins: $\alpha$-helices and $\beta$-sheets.
      * Identify which specific residues will inhabit the same face of an $\alpha$-helix or a $\beta$-sheet.
      * Master the identification of amphipathic structures, as these facilitate the determination of residue placement on structural faces.

The Hierarchical Layers of Protein Structure

• Primary ($1^{\circ}$) Structure: Defined as a specific, linear sequence of amino acids.

• Secondary ($2^{\circ}$) Structure: Refers to local folding patterns into repeating units.

• Tertiary ($3^{\circ}$) Structure: The comprehensive folding of a single polypeptide chain into a specific three-dimensional (3D) arrangement.

• Quaternary ($4^{\circ}$) Structure: The association and interaction of multiple polypeptide chains into a larger, functional protein complex.

The Relationship Between Structure and Function

• Necessity of Higher Order Structures: A simple primary sequence of amino acids in a disordered chain typically does not constitute a functional protein. Folding is an essential requirement for biological function.

• Protein Denaturation:
      * Denaturation occurs when a protein becomes unfolded, leading to a loss of function.
      * Typical causes of denaturation include changes in temperature, shifts in $pH$, or the addition of organic molecules.
      * Chemical Change during Denaturation: Noncovalent bonds are broken during the process, but the covalent bonds (including the peptide bonds of the primary structure) remain intact.

• Primary Structure as a Determinant of Folding:
      * The primary sequence is the major factor determining how a protein folds.
      * Amino acid stereochemistry is critical to secondary structure formation.
      * Influential R-group Properties:
          * Hydropathy (hydrophobicity/hydrophilicity).
          * Aromaticity.
          * Charge.
          * Size.
          * Coordination chemistry.
          * Chemical modifications.

Fundamental Determinants of Secondary Structure

• Local Folding: Secondary structure involves the N-terminus, C-terminus, and side chains folding into local repeating motifs.

• Example Case Study (Myoglobin): Myoglobin serves as an example of a protein consisting exclusively of $\alpha$-helical secondary structures, featuring a bound heme group.

• Physical and Chemical Constraints on $2^{\circ}$ Structure:
      1. Bond Integrity: Bond lengths and bond angles are fixed and cannot be distorted.
      2. Steric Hindrance: Atoms cannot infringe upon or overlap with each other's physical space.
      3. Peptide Bond Configuration: Peptide bonds must remain planar and usually exist in the trans configuration.
          * Proline Exception: Proline is the only residue that frequently deviates from this standard constraint.

• Rotational Freedom: Due to the planarity of the peptide bond, rotation is only possible around the two bonds adjacent to each central alpha carbon ($C\alpha$).

• Stabilization Forces: Folding is stabilized primarily by noncovalent bonds, with hydrogen bonding playing the most significant role.

Dihedral Angles and the Ramachandran Plot

• The Origin of Dihedral Angles: The specific geometry and stereochemistry of peptide bonds define the dihedral angles:
      * $\Phi$ (phi): Describes the rotation around the bond between the amide nitrogen ($N_{\text{amide}}$) and the alpha carbon ($C\alpha$).
      * $\Psi$ (psi): Describes the rotation around the bond between the alpha carbon ($C\alpha$) and the carbonyl carbon ($C_{\text{carbonyl}}$).

• Steric Restrictions: Interaction between atoms (sterics) significantly limits the allowed values for $\Phi$ and $\Psi$.

• The Ramachandran Plot:
      * Proposed in $1963$ by the Indian physicist Gopalasamudram Narayanan Ramachandran and his colleagues.
      * It is a visualization tool that maps allowed $\Phi$ and $\Psi$ angles.
      * Specific regions on the plot correspond to the structural motifs of $\alpha$-helices and $\beta$-sheets.

• Amino Acid Variations on the Plot:
      * Glycine (Gly; G): A nonpolar aliphatic amino acid. Due to its hydrogen side chain, it has much less steric hindrance and occupies more regions of the Ramachandran plot.
      * Proline (Pro; P): A cyclic amino acid. Its rigid ring structure severely restricts the allowed $\Phi$ and $\Psi$ angles compared to other residues.

• Specific Structural Regions on the Ramachandran Plot:
      * Antiparallel $\beta$ strands: Typically found near $\Phi = -135^{\circ}$ and $\Psi = +135^{\circ}$.
      * Parallel $\beta$ strands: Typically clustered near $\Phi = -120^{\circ}$ and $\Psi = +120^{\circ}$.
      * Polypeptide II helix: Located in the upper left quadrant near $\Phi = -75^{\circ}$ and $\Psi = +150^{\circ}$.
      * Right-handed $\alpha$ helix: Found in the lower left quadrant near $\Phi = -60^{\circ}$ and $\Psi = -45^{\circ}$.
      * $3_{10}$ helix: Found close to the right-handed $\alpha$ helix region.
      * Left-handed $\alpha$ helix: Found in the upper right quadrant near $\Phi = +60^{\circ}$ and $\Psi = +45^{\circ}$.

Characteristics of $\alpha$-Helices

• Hydrogen Bonding Pattern: $\alpha$-helices are stabilized by specific hydrogen bonds formed between the carbonyl oxygen ($C=O$) of residue "$i$" and the amide hydrogen ($NH$) of residue "$i + 4$".

• Orientation of Side Chains: In an $\alpha$-helix, side chains (R groups) radiate outward, away from the central helical axis. This is clearly visible when viewing the structure down the main chain axis.

Characteristics of $\beta$-Sheets

• Stabilization: $\beta$-sheets are stabilized by main chain hydrogen bonds occurring between individual $\beta$-strands.

• Constituents: A sheet must consist of two or more $\beta$-strands.

• Conformation: Each residue within a strand exists in an extended conformation. Dihedral angles $\Phi$ and $\Psi$ are typically both near $180^{\circ}$.

• Hydrogen Bonding Distance: The distance (in the primary sequence) between hydrogen bonding partners varies based on the arrangement of the sheet and the size of the intervening loops.

• Side Chain Orientation: Within a $\beta$-sheet, adjacent side chains are located on opposite faces (pointing in opposite directions) along the sheet.

• Strand Arrangements:
      1. Antiparallel Arrangement: Strands run in opposite directions relative to their N- and C-termini.
      2. Parallel Arrangement: Strands run in the same direction.

• Connectivity: $\beta$-strands are connected by turns or loops of variable sizes. These loops are influenced by the sheet's parallel or antiparallel nature and can even connect strands located on different polypeptide chains.

Periodicity and Amphipathic Patterns

• Pattern Recognition: Because secondary structures are periodic, diagnostic patterns may be visible in the primary amino acid sequence.

• Amino Acid Preferences: While certain amino acids prefer specific secondary structures, these preferences are considered "fuzzy" and are not absolute rules.

• Amphipathicity: A structure is amphipathic if it has a polar face and a non-polar face.

• $\alpha$-Helix Patterns:
      * Since $\alpha$-helices have approximately $3.6$ residues per turn, a pattern of polar/non-polar residues repeating every $3$ to $4$ positions indicates an amphipathic helix.
      * Example Sequences:
          * FKSEVNKK ($F, V$ are non-polar; others polar).
          * FLSEVAKK ($F, L, V, A$ are non-polar).
      * Visualizing the Helical Wheel: Plotting residues such as K, A, V, F, L, S, E, K on a wheel reveals separated polar and non-polar faces.

• $\beta$-Sheet Patterns:
      * Because side chains alternate faces in a $\beta$-strand, an amphipathic $\beta$-sheet is identified by a sequence where polar and non-polar residues alternate (every other residue).
      * Example Sequence: DLKVSF (where $D, K, S$ are polar/charged and $L, V, F$ are non-polar).