BMS1041 Biochemistry A: Protein Structure

• Study notes for BMS1041 Biochemistry A Conceptual Overview provided.
• These cover Week 1, focusing on protein structure and function.
• Presented by Dr. Maria Teresa Esposito, Senior Lecturer in Biochemistry.

Week 1 Outline:

• Amino acids, peptides, and proteins are covered in Lecture 1.
• The structure of proteins is covered in Lecture 2.
• Hemoglobin and Myoglobin protein function is covered in Lecture 3.

Lecture 1: Amino Acids, Peptides, and Proteins:

• Recall the general structure of an amino acid.
• Understand how amino acids are categorised
• Describe the formation of a peptide bond
• Understand the differences between the different levels of a protein structure

Essential Reading Material:

• Chapter 3 in "Lehninger Principles of Biochemistry" covers amino acids, peptides, and proteins.
• Sections 3.1 and 3.2 are particularly relevant.
• Chapter 3 of "Molecular Biology of the Cell" offers basic concept revision.

Structure of Amino Acids:

• Proteins are the most abundant biological macromolecules in all cells.
• They act as effectors for expressing genetic information.
• A common set of 20 amino acids constructs proteins.
• The general structure includes an α carbon, a carboxyl group (COO-), an amino group (NH3+), and a side chain (R group).

Classification of Amino Acid R-Groups:

• R groups differ in size and electric charge.
• R group variations impact the solubility of the amino acid in water.

Amino Acid Nomenclature:

• Amino acids have 3-letter abbreviations.
• They're classified as polar/uncharged, negative, positive, or nonpolar based on side chain properties.
• Examples include Alanine (nonpolar), Arginine (positive), Aspartic acid (negative), and Serine (uncharged polar).

Peptide Formation:

• Amino acids link covalently through a peptide bond.
• Combining two amino acids releases H₂O in a condensation reaction.
• The reverse reaction with water is hydrolysis.
• Two amino acids joined form a dipeptide, three form a tripeptide.
• Few amino acids form an oligopeptide.
• Many amino acids form a polypeptide or protein.
• Polypeptides have molecular weights under 10 kDa.

Oligopeptides and Polypeptides:

• Ser-Gly-Tyr-Ala-Leu Is shown as an example of a pentapeptide with five amino acid sequence.
• Amino acid sequence is the primary structure of a protein
• The sequence is written from the amino (N) terminus to the carboxyl (C) terminus.

Protein Quantification:

• Proteins with aromatic groups absorb UV light.
• The characteristic absorbance at 280 nm is used for protein characterization.
• The Lambert-Beer law applies.

Peptide Ionization:

• Peptides have one free N-terminus and one free C-terminus, plus ionizable R groups.
• Ionizable groups contribute to the protein's acid-base properties.
• Peptides have unique isoelectric points (pI).

Small Peptide Examples:

• Aspartame is an artificial sweetener.
• Oxytocin is a hormone with 9 amino acids.
• Amanitin is toxic from mushrooms.
• Antibiotics.

Protein Examples:

• Proteins can consist of either one polypeptide chain or multiple.
• Proteins with multiple subunits are multisubunit proteins.
• Subunits may or may not be identical.
• Identical subunits indicate an oligomeric protein and are called protomers.
• Hemoglobin consists of two identical α chains and two identical β chains.
• Insulin are linked covalently by disulfide bonds.

Conjugated Proteins:

• These proteins contain chemical components besides amino acids.
• The non-amino acid part is the prosthetic group.
• Lipoproteins contain lipids, and glycoproteins contain sugar groups.
• Metalloproteins contain specific metals.
• Prosthetic groups often play key biological roles.

Four Levels of Protein Structure:

• There are four levels of protein structure:
  • Primary is the amino acid sequence.
  • Secondary involves local folding into helices or sheets.
  • Tertiary is the 3D arrangement of a single protein.
  • Quaternary is the arrangement of multiple protein subunits.

Lecture 2: Continued Discussion on Four Levels of Protein Structure

• Primary Protein Structure: Sequence of a chain of amino acids.
• Secondary Protein Structure: Local folding of the polypeptide chain into helices or sheets.
• Tertiary Protein Structure: Three-dimensional folding pattern of a protein due to side chain interactions
• Quaternary Protein Structure: Protein consisting of more than one amino acid chain.

Learning Outcomes For Lecture 2

• Describe the arrangement of amino acids and bonding in alpha- helices and beta- sheets
• Explain the different types of bonds that determine the tertiary structure of a protein.
• Understand the terms folding, unfolding, misfolding and denaturation.
• Understand what a ligand is
• Explain the term binding site
• Understand the association and disassociation are opposites, what the dissociation constant K, is and that a low K₁ means a protein binds a ligand tightly.

Additional Reading

• Lehninger, Principles of Biochemistry covers the structure of three-dimensional proteins in chapter 4.
• 4.1-4.4 are relevant to protein structures, denaturation and folding.

Primary Structure Details:

• It consists of a linear amino acid sequence created by peptide bonds.
• Primary structure forms during translation.
• It can be modified by post-translational changes.

Modified Amino Acids in Proteins:

• Some proteins contain residues from modified common ones post-synthesis.
• Examples include 4-hydroxyl proline, y carboxy glutamate, and desmosine
• Can also modify by addition of phosphoryl, methyl, acetyl, adenylyl, ADP-ribosyl to their amino acids.
• These modifications are reversible

3D Protein Conformation:

• The covalent backbone allows numerous potential conformations.
• Proteins adopt a specific three-dimensional structure related to function.
• Conformations are stabilized by both weak non-covalent forces and covalent bonds in the backbone.

Covalent Peptide Bond Structure:

• Alpha carbons of residues are separated by three arranged covalent bonds
• Six atoms of a peptide group lie in a single plane because of X ray diffraction studies.
• The nitrogen is partially positive and the oxygen is partial negative, creating a small dipole
• The peptide bond is rigid, planar and cannot rotate freely

Peptide Bond Characteristics:

• Peptide conformation is defined by Φ (Phi), Ψ (Psi), and Ω (Omega) dihedral angles.
• Angles reflect rotation about repeating bonds.
• A dihedral angle is the intersection between two planes.

Solving Protein Structure:

• Frederick Sanger sequenced amino acid residues of hormone insulin.
• Used an Edman degradation technique which won him a Nobel Prize for Chemistry in 1958
• Dorothy Hodgkin used X-ray crystallography to solve the structure of penicillin, vitamin B12, and insulin.
• Won the Nobel Prize for Chemistry in 1964 for her work

Protein Sequencing Advancements:

• Protein sequencing is achieved by matrix assisted laser desorption/ionization mass spectrometry or MALDI-MS.

Secondary Structure:

• Secondary structure involves protein chains in helices, sheets, and turns.
• Amino acids are arranged into either a helices or ẞ sheet.
• Hydrogen bonds are present
• Can form fibrous or globular proteins

Alpha Helix Details:

• It is generated when a single chain twists.
• Hydrogen bonds form between every fourth peptide bond.
• Hydrogen forms between C=O of and N-H
• N-H groups point up and C=O points down, this gives it polarity
• Right-handed helices are most common.
• The R groups are located outside the helix, and there is a turn every 3.6 residues.
• Alpha helixes are abundant in cell membranes.

Alpha Helix Destabilizing Factors:

• Charged amino acids (Asp, Glu, Lys, Arg) destabilize structure.
• Certain polar, uncharged amino acids (Asn, Gln, Ser, Thr, Cys).
• Proline causes kinks.
• Glycine is too flexible.
• Alanine has the greatest tendency to form a helices.

Beta Strands and Beta Sheets:

• Adjacent peptide chains in ẞ sheets run in opposite (antiparallel) or parallel directions.
• Hydrogen bonds connect peptides in different strands.
• Antiparallel sheets have stronger hydrogen bonding.
• Flexible loops and turns (Pro, Thr, Ser, Gly) link secondary structure regions.

Beta Turns:

• Common in globular proteins
• Structure allows some amino acid residues to form turns and loops and assume a very compact structure.
• Beta turns are the connecting elements that link successive runs of a helix or ẞ conformation.
• Gly and Pro residues often are in beta turns.

Circular Dichroism Spectroscopy:

• Structural asymmetry in molecules causes differences in polarized light absorption.
• This difference is measured through circular dichroism spectroscopy.
• CD spectra vary for a and ẞ helices and unstructured proteins.
• CD spectra provide estimates of common secondary structures.

Tertiary Structure:

• 3D conformation is maintained by disulfide bonds, salt bridges, and hydrogen bonds.
• Examples: insulin, hemoglobin, myoglobin, and hormones.
• Proteins may be fibrous or globular in shape.

Fibrous Proteins:

• Fibrous proteins: such as polypeptide chains that connect arrange on long strands or sheets.
• Globular proteins: polypeptide chains arranged into a spherical or globular shape.
• Membrane proteins: have polypeptide chains that are embedded in hydrophobic lipid membranes.

Protein Stabilization:

• The hydrophobic effect dominates weak stabilizing forces.
• Hydrogen bonds, van der Waals interactions, and electrostatic interactions also contribute.

Fibrous protein examples:

• Keratin provides strength/flexibility with fundamental structure.
• Consists of two wrapped alpha helices.
• Stabilizing cross-links in quaternary structure with disulfide bonds.
• Collagen is fundamental for connective strength.
• Consists of 3 entwined alpha helices and related to brittle bone issues.
• Fibroin used produces silk in beta conformation.
• Does not stretches for extended use, kept together by weak interaction.

Globular Proteins:

• Globular proteins roughly spherical with secondary structures.
• Often soluble, with hydrophobic.
• Enzymes, regulatory and other examples: hemoglobin, cytokines actin and myosin.
• Composed of different motifs folded together.

Myoglobin:

• Function is for oxygen diffusion to rapidly contracting muscle.
• The backbone consists of a helices and has four parts of water. Also a single protein group.
• It consists of a single polypeptide chain with a single iron protoporphyrin (heme) group.

Structures of the Heme:

• Iron protoporphyrin with bounds ferrous 2+
• Present in myoglobin and hemoglobin

Myoglobin Binding to Oxygen:

• Proteins interacts with molecules
• Binding that complement protein and binding to ligands
• Binding can cause conformational changes , like an induce fit
• It must be bound able to release to binding sites
• It's chemically alter to enzymes

Association and Dissociation:

• Association is protein binds, disassociation is ligand released.
• A high is for a protein ligand to use.
• Relationship where its is known the association equation
• the Ka in disruption to disassociation for that

Fractions of Usable Binding Sites:

• Some will occupy to each protein ligand sites
• Fraction to some

Oxygen Binding to Heme:

• Oxygen binding from that prosthetic
• Is from hemme dissociation cuvere
• Some myoglobin to use

Lecture 3: Myoglobin and Hemoglobin - Learning outcomes

• Explain the term allosteric.
• Compare the structures of myoglobin and haemoglobin.
• Explain how haemoglobin binds and releases O2, CO2 and H+.
• Interpret oxygen binding curves.

Additional Reading Material

• Lehninger Principles of Biochemistry - Protein function chapter 5
• 5.1 - reversible binding of protein

Quaternary Structure

• Some proteins contain two separated polypeptide chains.
• Has quaternary structure with multiple that turn to functional
• Regulatory - others having regulatory with function

General Hemoglobin Information:

• Made of two a and two b polypeptidees
• Subunits related to myoglobin and carry oxygen in blood
• Transitions between two confirmations

Oxygen Cooperativity:

• The more O2 that Hb bind
• Each molcule can hold a Max if 4 O2

Allosteric Cooperativity:

• Allosteric cooperativity happens as more oxygen binds.
• CO2 as low to is CO
• Hb transitions form low -> high affinity
• Hemoglobin efficient when released and oxygen.
• CO is Haem 250 more to ready than O2

Hemoglobin Transports H:

• CO
• CO 2 at N

Oxygen in Low PH:

1. • low H& HCO
2. → Low and hH+ to hemoglobiin

2, 3 Beta Glyerophosphate:

• BPG regulate to bind a hemodbilin
• PO2 Low altitude
• Higher altitude + POG low blood -> O2 Affinity

BPG Decreases Oxygen Infinity

• Low - slightly is high
• Low/high [BPG]: