Peptides, Proteins, Protein Functions
Course Learning Objectives
BIOC13: Hypothesize protein structure and function based on sequence characteristics.
BIOC14: Assess structural and environmental changes in protein structure.
Module Learning Objectives (MLOs)
13.1: Predict experimental results for reactions of proteins or peptides with various proteases or chemical treatments.
14.1: Describe the factors that allow each protein to maintain a specific shape and perform a specific function.
14.2: Identify various structural elements and motifs within a protein.
14.3: Describe factors affecting protein folding and denaturation.
14.4: Describe common protein pharmaceuticals and protein folding diseases.
Understanding Peptides
Peptides: Chains of amino acids linked together by peptide bonds, typically formed through dehydration reactions (releasing a water molecule).
Peptide bond: An amide linkage between amino acids.
Naming of Peptides: Peptides are always named from the N-terminus to the C-terminus.
Polyamphoteric: A polypeptide that possesses one or more amino acids with ionizable side chains, leading to multiple pKa values.
Impact of pH on Protein Shape: Shifts in pH can significantly affect the shape of a peptide as these side chains determine peptide conformation.
Bond Angles in Peptide Bonds
Peptide bonds exhibit partial double bond character due to overlap.
Resonance structure: Peptide bonds are not static and can exist in two forms:
Trans: Typically favored due to steric hindrance.
Cis: Possible but less favored in proline.
Cleavage of Peptides
1. Enzymatic Cleavage
Enzymes: Necessary to hydrolyze a polypeptide due to the slow rate of uncatalyzed hydrolysis at physiological pH.
Digestive enzymes (proteases): Cleave peptide bonds at specific sites:
Trypsin: Cuts after positive amino acids (Arg and Lys) unless proline follows.
Chymotrypsin: Cuts after large hydrophobic amino acids (Phe, Trp, Tyr) unless proline is before them.
Pepsin: Cuts before amino acids (Leu, Phe, Trp, Tyr) unless proline is the preceding amino acid.
2. Chemical Cleavage
Cyanogen bromide (CNBr): Cleaves after methionine amino acids.
Edman Degradation: Chemically cleaves amino acids from the N-terminus one at a time, independent of the amino acid present; used for protein sequencing.
Combining Cleavage Methods
Utilizing these cleavage methods helps digest proteins into shorter fragments for easier sequencing.
Some drugs exploit protease mechanisms — e.g., protease inhibitors prevent the maturation of retroviruses by blocking specific cleavage steps.
Understanding Protein Structure
Proteins: Defined polypeptides with a specific sequence that dictates:
Specific folding patterns.
Overall structure, enabling them to perform unique functions.
Primary Structure: The amino acid sequence dictated by the genetic code.
Rotational Degrees of Freedom: Associated with conformational angles;
(psi (Ψ)) and (phi (φ)).
The Role of Sequences in Folding
Specific amino acid combinations exhibit minimal steric strain, reproducibly forming expected structures revealed by Ramachandran plots.
Levels of Protein Structure
Primary Structure
Defined as the amino acid sequence alone.
Secondary Structure
Convention: Local folding that can be predicted.
Alpha Helix Characteristics:
Found when amino acid angles Ψ and φ are approximately -50° and -60°.
Contains 3.6 residues per turn, stabilized by hydrogen bonds between C' = O of residue n and N-H of residue n+4.
Exhibits a distinct dipole moment with a negative charge at the C-terminus and a positive charge at the N-terminus.
Beta-Pleated Sheets
Aligned in either:
Parallel
Antiparallel
Typically contain 5-10 residues with ψ and φ angles in the upper left quadrant of Ramachandran plots.
Stabilized by hydrogen bonds:
Form between C' = O groups on one strand and N-H groups on another.
Pleated structure with atoms above and below the plane.
Tertiary Structure
Represents the overall fold of a protein:
Globular proteins define inside and outside.
Beta sheets may form barrel structures, with polypeptide chains demonstrating flexibility in turns.
Quaternary Structure
Organization of multiple subunits or domains; not all proteins achieve this level of structure.
Example: Hemoglobin requires multiple subunits for functional operation.
Protein Folding
Information for three-dimensional structure is predominantly contained in the amino acid sequence.
Protein Denaturation: Influenced by various factors, including changes in pH, which modify hydrogen bonding:
is positive when unfolding is unlikely, therefore proteins remain folded.
Factors Influencing Protein Stability
Temperature: Increased temperature leads to protein unfolding.
Chemical Denaturation: Increased urea concentrations destabilize proteins.
Ionic Strength: Altering ionic strength affects protein solubility.
Salting In: Increases solubility of proteins but may lead to instability.
Salting Out: Increases protein stability while decreasing solubility.
Oxidation/Reduction and Effects on Protein Structure
Cysteine residues allow proteins to exist in the oxidized state (S-S bonds) or reduced state (-SH).
Benefits of Proper Protein Folding: Maximizes interactions that stabilize the protein structure (e.g., hydrogen bonds, buried nonpolar groups).
Renaturation Post-Denaturation
Some proteins can be renatured by correctly adjusting environmental conditions.
Chaperonins: Assist in protein folding during biosynthesis, one example being Gro-EL.
Protein Pharmaceuticals
More than 100 FDA-approved protein drugs, primarily recombinant proteins, include the following classes:
Vaccines
Peptide medications
Blood products
Recombinant therapeutic proteins
Issues include denaturation, aggregation, and proteolysis by digestive enzymes.
Protein Modifications Impacting Drug Stability
Modifications may include deamidation, disulfide exchange, glycosylation, and phosphorylation.
Protective delivery methods, including freeze-drying and transdermal patches, are utilized to prevent instability.
Prion Diseases
Characterized by misfolded proteins (PrP):
Animal Prion Diseases
Bovine Spongiform Encephalopathy (BSE): Infectious strain from contaminated feed.
Chronic Wasting Disease (CWD)
Scrapie: Spreads via contaminated nerve tissue.
Transition from normal PrPC (soluble and α-helix) to misfolded PrPSc (insoluble and β-sheet).
Human Prion Diseases
Creutzfeldt-Jakob Disease (CJD): Includes sporadic, familial, and contamination cases.
Notable symptoms: A range from social withdrawal to coma.
Kuru: Infectious, transmitted through consuming human brain tissue.
Functional Aspects of Proteins
1. Structural Roles
Fibrous Proteins: Elongated forms serve structural roles in cells.
α-Keratins: Predominantly α-helical, integral to hair and skin.
Fibroin: Composed of antiparallel β-sheets.
Collagen: Triplet helix made of left-handed helices, predominantly glycine residues; vitamin C is crucial for proline hydroxylation.
2. Contractile Roles
Actin-Myosin Complex: Utilizes ATP hydrolysis for energy during contraction.
3. Defense Proteins
Antibodies: Respond to foreign antigens by binding and neutralizing them through complex processes involving T-cells and B lymphocytes.
4. Transport Proteins
Hemoglobin: Integral in oxygen transport, further details anticipated in subsequent sections.
5. Signaling Mechanisms
Utilizes pathways such as adenylate cyclase for transducing hormonal signals into cellular responses, including the phosphorylation of metabolic enzymes.
Conclusion of the Notes
The structure and function of proteins are deeply intertwined, dictating cellular and physiological roles essential for life. Understanding the nuances of their formation, stability, and reactivity can inform therapeutic applications and disease management.