Protein Chemistry Lecture Notes

MBMB 208 - Protein Chemistry - Lecture 1 - Introduction

Course Overview

  • Proteins are essential macromolecules that play critical roles in the normal functioning of living organisms.
  • They are products of DNA translation, composed of amino acids linked by peptide bonds.
  • Proteins are biological laborers, crucial for the structure, function, and regulation of cells and tissues.
  • Proper protein folding (3D structure) is essential for function; misfolding due to changes in amino acid sequence can lead to dysfunction and disease.
  • Protein chemistry is central to biochemistry and related fields like immunology, biophysics, enzymology, and endocrinology.
  • This course equips students with protein chemistry fundamentals.
  • Topics include protein structure, protein-ligand interactions, chemical bonds/reactions in protein/protein and protein-ligand interactions, and mechanisms of enzyme action.

Learning Outcomes

  • Describe the machinery involved in protein translation.
  • Distinguish between primary, secondary, tertiary, and quaternary structures.
  • Describe the steps involved in determining amino acid composition and sequence of a protein.
  • Work out complete primary structures.
  • Describe the peptide bond and its effect on secondary structure.
  • Describe different secondary structures (regular and irregular).
  • Make a structural distinction between fibrous proteins and globular proteins.
  • Discuss common examples of proteins with tertiary and quaternary structures.
  • Discuss protein-ligand interactions.
  • Describe and compare the structures of myoglobin and hemoglobin.
  • Explain the mechanism of action of hemoglobin as an oxygen transport protein.
  • Distinguish between cooperative and noncooperative ligand-binding.
  • Discuss allostery and the different models of allosteric interactions.
  • Discuss the various mechanisms of enzyme action with examples.

Lecture Outline

  • The central dogma of molecular biology
  • DNA replication and transcription
  • mRNA processing
  • The genetic code and tRNA
  • Ribosomes and rRNA
  • Major classification of amino acids

The Central Dogma of Molecular Biology

  • The central dogma outlines the flow of genetic information:
    • DNA is replicated.
    • DNA is transcribed into RNA.
    • RNA is translated into protein.
  • Reverse transcription also occurs, where RNA is converted back into DNA.

Introduction to Amino Acids (AA)

  • Amino acids are the monomer units from which polypeptide chains of proteins are synthesized.
  • They are organic compounds containing amine ($-NH_2$) and carboxyl ($-COOH$) groups.
  • Common amino acids are $\alpha$-amino acids, with the amino group attached to the $\alpha$-carbon (the carbon next to the carboxyl group).
  • Proline is an exception, possessing a secondary amino group ($-NH-$), but is still considered an $\alpha$-amino acid.

Amino Acid Structure

  • Tetrahedral carbon: A carbon atom with four attachments and bond angles of approximately 109.5o109.5^o. The 20 standard amino acids differ in their side chains (R groups).
  • The side chain (R group) gives each amino acid its unique identity.

Classification of Amino Acids

  • Amino acids are classified into five main classes based on the properties of the R group:
    • Nonpolar amino acids (9 amino acids):
      • Alanine, Valine, Leucine, and Isoleucine (aliphatic hydrocarbon side chains).
      • Proline (cyclic pyrrolidine).
      • Methionine (thioether side chain, one of the two sulfur-containing amino acids).
      • Phenylalanine (phenyl moiety) and tryptophan (indole group) are aromatic amino acids.
      • Tryptophan is sometimes considered a borderline member because it can interact favorably with water via the N-H moiety of the indole ring.
      • Glycine can also be considered as polar uncharged
    • Polar, Uncharged Amino Acids (6 amino acids):
      • All, except glycine, contain R groups that can:
        • Form hydrogen bonds with water.
        • Play nucleophilic roles in enzyme reactions.
      • These amino acids are more soluble in water than nonpolar amino acids.
      • Asparagine and Glutamine (amide group).
      • Tyrosine, Threonine, and Serine (hydroxyl group).
      • Cysteine (sulfhydryl group).
    • Charged Polar Amino acids (5 amino acids):
      • Side chains are positively or negatively charged at physiological pH values.
      • Basic amino acids are positively charged (+):
        • Histidine (imidazole group).
        • Arginine (guanidino group).
        • Lysine (protonated alkyl amino group).
      • Acidic amino acids are negatively charged above pH 3:
        • Aspartic acid and glutamic acid (R groups contain a carboxyl group).

Other Ways to Classify Amino Acids

  • Hydrophobic: Alanine, Valine, Isoleucine, Leucine, Phenylalanine, Proline.
  • Hydrophilic: Arginine, Asparagine, Aspartic acid, Cysteine, Glutamic acid, Glutamine, Histidine, Lysine, Serine, Threonine.
  • Amphipathic: Methionine, Tryptophan, Tyrosine, Glycine

Structures of Amino Acids

  • Nonpolar (hydrophobic): Leucine, Proline, Alanine, Valine, Methionine, Phenylalanine, Tryptophan, Isoleucine
  • Polar, uncharged: Glycine, Serine, Asparagine, Glutamine.
  • Acidic: Aspartic acid, Glutamic acid.
  • Basic: Lysine, Arginine, Histidine

Nutritional Classification of Amino Acids

  • Essential amino acids: The body cannot synthesize them in sufficient quantities, so they must be obtained from the diet. Examples: Valine, isoleucine, lysine, leucine, methionine, tryptophan, threonine, and phenylalanine.
  • Semi-essential amino acids: Depending on individual needs and physiological conditions (e.g., rapid growth or illness), some non-essential amino acids may become semi-essential. Examples: Arginine and Histidine.
  • Non-essential amino acids: The body can produce these internally based on dietary requirements. Examples: Alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, proline, serine, and tyrosine.

Symbols/Abbreviations for Amino Acids

  • Standard three-letter and one-letter abbreviations are used to represent amino acids.

Class Activity

  • Identify the amino acids in your name
  • Draw a generic amino acid and identify its $\alpha$ carbon and its substituents.
  • Draw Fischer projections for the two enantiomers of Alanine, Leucine, Valine.
  • Draw the structures of the 20 standard amino acids and give their one- and three-letter abbreviations.
  • Classify the 20 standard amino acids by polarity, structure, type of functional group, and acid–base properties.

The Acid-Base Properties of Amino Acids

  • Common amino acids are weak polyprotic acids. The degree of dissociation depends on the pH of the medium.
  • Amino acids contain at least two dissociable hydrogens.
  • Amino acids can act as both acids and bases (amphoteric) due to the amino and carboxyl groups.
  • Zwitterions/Dipolar ions: Compounds with separate parts that are positively and negatively charged but with no overall electrical charge.
Amino Acid Forms at Different pH Levels
  • Low pH: Cationic form, net charge +1
  • Neutral pH: Zwitterion form, net charge 0
  • High pH: Anionic form, net charge -1
Protonic Equilibria of Aspartic Acid
  • In strong acid (below pH 1); net charge = +1
  • Around pH 3; net charge = 0
  • Around pH 6-8; net charge = -1
  • In strong alkali (above pH 11); net charge = -2

pKa and Isoelectric Point (pI)

  • Acid strengths of weak acids are expressed as their pKapK_a.
  • Isoelectric point (pI): The pH at which the total charge of the amino acid (peptide or protein) is zero.
  • pI=(pK<em>a1+pK</em>a2)/2pI = (pK<em>{a1} + pK</em>{a2}) / 2

pKa Values of the 20 Common Amino Acids

A table is provided with the pK<em>a1pK<em>{a1}, pK</em>a2pK</em>{a2}, and pKaRpK_{aR} values for the 20 common amino acids.

Titration of Glycine

  • Glycine exists in different forms depending on pH.
  • pI=(pK<em>1+pK</em>2)/2pI = (pK<em>1 + pK</em>2) / 2
  • pI=(2.3+9.6)/2pI = (2.3 + 9.6) / 2
  • pI=11.9/2pI = 11.9 / 2
  • pI=5.95pI = 5.95

Isoelectric Point Calculations

  • For polyfunctional acids, pI is the pH midway between the pKapK_a values on either side of the isoionic species.
    • Example for aspartic acid: pI=(pK<em>a1+pK</em>a2)/2=(2.09+3.96)/2=3.02pI = (pK<em>{a1} + pK</em>{a2}) / 2 = (2.09 + 3.96) / 2 = 3.02
    • Example for lysine: pI=(pK<em>a2+pK</em>a3)/2=(8.94+10.53)/2=9.73pI = (pK<em>{a2} + pK</em>{a3}) / 2 = (8.94 + 10.53) / 2 = 9.73

Peptide Bond Formation

  • Amino acids link via their amino and carboxyl groups, forming a covalent amide bond (peptide bond) by releasing water.
  • This reaction requires energy.
  • In water, the reverse reaction (hydrolysis) is favored.
  • Repeated peptide bond formation creates polypeptides and proteins.

Peptide Length

  • 2 AA joined by a peptide bond: dipeptide
  • 3 AA: tripeptide
  • 4 AA: tetrapeptide
  • 5 AA: pentapeptide
  • Few AA: oligopeptide
  • Many AA: polypeptide

Naming of Peptides

  • Aminoacyl residues are named by replacing the -ate or -ine suffixes with -yl (e.g., alanyl, aspartyl, tyrosyl).
  • Peptides are named as derivatives of the carboxyl-terminal aminoacyl residue. For example, Lys-Leu-Tyr-Gln is named lysyl-leucyl-tyrosyl-glutamine. The -ine ending on glutamine indicates that its $\alpha$-carboxyl group is not participating in peptide bond formation.
Peptide Bond Formation Depiction
  • Amino acid 1 + Amino acid 2 → Dipeptide + H2OH_2O

Proteins

  • Proteins with one polypeptide chain are monomeric proteins.
  • Proteins with more than one polypeptide chain are multimeric proteins.
  • Multimeric proteins may be homomultimeric (containing only one kind of polypeptide) or heteromultimeric (composed of several different kinds of polypeptide chains).

Protein Structure

  • Primary structure: Amino acid sequence.
  • Secondary structure: Localized structures like $\alpha$-helices and $\beta$-sheets formed by hydrogen bonds between amino acids.
  • Tertiary structure: The overall three-dimensional structure of a single polypeptide chain.
  • Quaternary structure: The arrangement of multiple polypeptide subunits in a multi-subunit protein.

Reading Assignment

  • Protein separation and purification
  • Quantification or estimation of amino acid sequence (Electrophoresis; SDS electrophoresis, Isoelectric focusing, Mass spectrometry, Matrix-Assisted Laser Desorption/ Ionization Mass Spectrometry, or MALDI MS_ mass-to-charge ratio, m/z etc)
  • NB//: Proteins functions by the amino acid sequence

References

  • Voet, D, Voet, J and Pratt, C.W. Fundamental of Biochemistry_ Life at the molecular level 5th edition
  • Garrett RH, Grisham CM. (2012) Biochemistry. 7th Ed, Cengage
  • Nelson DL & Cox MM (2012). Lehninger Principles of Biochemistry, 6th edn. WH Freeman and Company, New York.
  • Rodwell V, Bender D, Botham KM., Kennelly PJ, Weil PA. Harper’s Illustrated Biochemistry. 30th Ed, McGraw-Hill Education, Lange