W26N Proteins

MPharm Programme: Amino Acids & Peptides

• Bruice's "Organic Chemistry" and Berg, Tymoczko, Stryer's "Biochemistry" are recommended readings.
• Nelson and Cox's "Lehninger Principles of Biochemistry" and Fersht's "Structure and Mechanism in Protein Science" are additional resources.

Proteins in Nature

• Enzymes, hormones, storage proteins, transport proteins, structural proteins, protective proteins, contractile proteins and toxic proteins represent some functions of proteins

Amino Acids

• Proteins and various crucial biomolecules derive from amino acids.
• Alpha-amino acids are nature's most prevalent amino acids.
• The generalized structure of an alpha-amino acid shows an amine group, carboxylic acid, side chain and a carbon atom.

Amino Acids: Chirality and Forms

• Glycine stands out as the only achiral amino acid among the 20 common ones.
• The remaining amino acids in nature exist in the L-optical form.
• Amino acids encompass both acidic and basic components.
• Nonionic and Zwitterionic forms are types of amino acids.

20 Amino Acids (i): Nonpolar, Aliphatic R Groups

• Glycine, alanine, proline, valine are amino acids with nonpolar, aliphatic R groups.
• Leucine, isoleucine, and methionine are additional examples.

20 Amino Acids (ii): Polar, Uncharged R Groups

• Serine, threonine, and cysteine compose the polar, uncharged R group amino acids.
• Asparagine and glutamine are additional examples.

20 Amino Acids (iii): Aromatic R Groups

• Phenylalanine, tyrosine, and tryptophan represent aromatic R group amino acids.
• These are signaling molecules

20 Amino Acids (iv): Positively Charged R Groups

• Lysine, arginine, and histidine characterize positively charged R group amino acids.
• Often in the active site of enzymes

20 Amino Acids (v): Negatively Charged R Groups

• Aspartate and glutamate represent negatively charged R group amino acids.
• Occur at a physiological pH

Amino Acid Composition and Isoelectric Point

• Every amino acid possesses its own isoelectric point.
• For neutral amino acid side chains, pl is 1/2 (pKaCOOH+pKaNH3+).
• In acidic side chains, pl equals 1/2 (pKaCOOH+pKar).
• Basic amino acid side chains have a pl of 1/2 (pKar+pKaNH3+).

Amide Formation: Carboxylic Acids and Amines

• Carboxylic acids and amines participate in condensation to produce amides, releasing water.

Oligopeptides, Polypeptides, and Proteins

• Aspartame, comprising two amino acids, exemplifies a dipeptide.
• Oligopeptides, presenting 2-10 amino acid residues, use suffixes such as di, tri, tetra, penta, hexa, hepta, octa, nona, and deca.
• Polypeptides are known to contain between 10 to 50 residues with an Mw under 5,000.
• Generally, proteins include 50-400 residues, boasting a Mw of 5,000-400,000.

Peptide Bond Characteristics

• Peptide bonds exhibit strong resonance.
• Delocalization energy stabilizes most peptides by 75-90 kJ/mol.
• Peptides favor a planar, trans (anti / Z) conformation.

Peptide Chain Examples

• Alanylglutamyl-glycyllysine demonstrates one example of a tetrapeptide, commencing with N-terminal AA.
• In a peptide chain, its charged groups at pH7 are represented by a red color.
• Serylglycyltyrosylalanylleucine exhibits one case of pentapeptide.

Cysteine and Disulfide Bond Formation

• Cysteines can be crosslinked through disulfide bonds.

Antioxidant Glutathione

• Glutathione, comprised of Glu-Cys-Gly, links via the sidechain Glu acid.

Medically Important Small Peptides

• Leucine enkephalin / Methionine enkephalin act as pain control.
• Bradykinin, Vasopressin and Oxytocin are other medically important peptides

Protein Structure and Analysis: Insulin

• In protein structure, insulin emerges as a notable example, crafted from dual amino acid chains interconnected via disulphide bonds.

Protein Purification

• Polyacrylamide gel electrophoresis (PAGE) contributes to analysis.
• Agarose is used for DNA, large proteins and enzyme complexes.
• Mass, shape, and charge determine movement.
• Sodium dodecyl sulphate (SDS) can denature proteins into uniform rods.

pH Electrophoresis and Protein Analysis

• pH electrophoresis aids in determining protein isoelectric points.

Amino Acids: Summary

• Amino acids (AA), numbering twenty common types, form structures and functions.
• They link via peptide bonds, forming amides.
• Short AA chains form oligopeptides, while larger ones form polypeptides and large proteins.
• AA sequences use amino terminus naming conventions.
• Besides peptide linkages, AA can bond via disulphide bonds.
• The isoelectric point impacts amino acid, peptide, and protein characterization and separation.

Protein Primary Structure

• Protein primary structure relates to amino acid sequence, demonstrated in peptides like Ser-Gly-Tyr-Ala-Leu.

Protein Structure: Disulphide Bonds in Hair

• Disulphide bonds greatly influence hair structure; they can be reduced, curled, and oxidized.

Secondary Protein Structure: Hydrogen Bonding

• Secondary structure relies on hydrogen bonding, elucidated by X-ray diffraction of fibrous proteins and small peptides.
• The α-helix and β-sheet emerge as major types.
• a-keratin and alpha-helices relate to hair and are always right handed

B-Keratin and Linking Structures

• β-keratin, found in silk, consists of β-sheets in parallel or antiparallel orientations.
• Other secondary structures include β barrels and twisted β sheets.
• Linking together is another aspect of Protein structure involving beta turns

Complex Proteins and Crystallization

• Crystallizing proteins can be challenging; RNaseA was effectively crystallized in space.
• NMR offers a useful alternative to X-ray techniques.
• Proline isomers pose issues in protein conformation.

Protein Conformation and Ramachandran Plots

• Classifying peptide conformations helps understand structure leading to the creation of ramachandran plots
• Ramachandran Plot showcases L-Ala which is a non-symmetric
• Ramachandran diagrams for rabbit pyruvate kinase, highlight the presence of Gly with conformational freedom over other amino acids

Protein Tertiary Structure: Packing and Interactions

• Myoglobin contains multiple α-helices.
• These elements combine into a compact globular shape; interactions include Van der Waals forces, hydrophobic effects, electrostatic forces, and hydrogen bonds.

Common Protein Tertiary Structures

• Serum albumin, ferritin, glycosyltransferases of the superhelical fold, engrained homeodomain.

More Common Protein Tertiary Structures

• a-Amylase inhibitor, trimeric LpxA-like enzymes, Hemopexin-like domain, a-D-Mannose, Immunoglobulin, Immunoglobulin-like, pilin, thymidylate, green fluorescent protein and Ubiquitin-conjugating enzyme make up more examples of protein tertiary structures

Protein Quaternary Structure and Interactions

• Proteins may spontaneously refold.
• Hemoglobin comprises 4 myoglobin-like sections for oxygen transport.
• Strands are held by non covalent forces such as hydrophobic side chains, tertiary helica structures

Transmissible Spongiform Encephalopathies (TSEs)

• Spongiform encephalopathies (prion diseases) are triggered by protein agents, exemplified by BSE, Scrapie, and Creutzfeldt-Jakob Disease.
• This lacks encoding from nucleic acids
• Misfolding relates to these encephalopathies

Protein/Peptide Purity

• Lysosomes and more are assembled with multiple structures.

Protein Sorting and Secretion: An Overview

• Proteins are active everywhere

Protein Sorting

• mRNA in the nucleus is transcribed and transported to the cytosol for translation by ribosomes.
• non-secretory and secretory pathways dictate protein destiny/processes

Secretory Pathway and Protein Sorting

• Eukaryotic cells use the same methods for synthesizing and secreted proteins and proteins that are found in the luminal space and in membranes by following 3 steps.

Signal Sequences and Protein Targeting

• Proteins for sorting undergo N-terminal signal sequencing-directing ribosomes to the endoplasmic reticulum (ER).
• The signal recognition particle (SRP) delivers them to the ER.

Protein Secretion: Co-translational Translocation

• Signal sequences on mRNA are translated then bound by signal recognition particle (SRP). This requires GTP
• SRP targets to the ribosome on the ER membrane. Signal peptidase cleaves
• Protein is folded into the ER and released into the cytosol.

Protein Sorting

• Signal sequences direct protein transit and integration.
• Hydrophobic elements determine destination.

The Cell Membrane And Protein Targeting Sequences

• The table demonstrates some known sorting singles that transport proteins to specific transport vesicles

Secretory and Endocytic Pathways

• The orientation is critical
• The protein remains until a domain is encountered
• There is single translocation

Secretory Pathway Overview and Vesicle Dynamics

• Vesicles play an essential protein process. Coated such as COPII and COPI exist with differing transports

Molecular Mechanisms in Vesicle Transport

• A vesicle fuses with a target membrane thanks to Snares
• Copll and Sec 12 relate to endoplasmic reticulum

Endocytosis

• The endocytic pathway, essential for bringing molecules into the cell, helps control many key functions using clathrin-coated complex

Protein Modifications

• Membrane and soluble secretory proteins, are the main process
• protein modification, for carbohydrate addition and structural maintenance, is critical to the in vivo protein folding and packaging processes