Understanding Protein Structure and Function

• The PH1123 lectures introduce protein and enzyme biochemistry
• Contact Dr. Alex White at [email protected]

What are Proteins?

• Proteins were discovered in 1838; the term "Proteios" means first or primary
• Proteins are abundant in biological systems, with each having a specific but initially unknown function
• Proteins are polymers made of 20 distinct amino acid monomers
• Each amino acid is identified by a three- or one-letter code
• Amino acids are classified based on their side chains (R) into seven classes: Aliphatic, Aromatic, Alcoholic, Sulfur Containing, Acidic, Basic, or Amide.
• The side chain determines the chemical property of the protein

Peptide Bonds and Protein Structure

• A peptide bond is a result of bonding between the carboxylic acid and amino groups of two adjacent amino acids with protruding R groups.
• The continuous peptide backbone folds into a specific three-dimensional shape
• Each protein has a unique and complex structure
• Visualizing the backbone alone facilitates easier studying
• An enzyme in spacefill illustrates all atoms (except hydrogen)
• This can demonstrate the compact, spherical nature of enzymes.

Functions of Proteins

• Proteins' complex structures facilitate diverse functions
• Catalysis: Enzymes are proteins facilitating chemical reactions; several thousand known.
• Storage and Transport: Ferritin stores/transports iron; Haemoglobin transports oxygen
• Mechanical Support/Shape: Collagen, abundant in vertebrates, is a supportive tissue component
• Decoding Information/Gene Expression: RNA polymerase synthesizes RNA (directed by DNA)
• Specialist Functions: Immunoglobins (antibodies) and some hormones (e.g., insulin)
• Proteins control almost every function in the human organism

Aliphatic Amino Acids

• Aliphatic amino acids include Glycine, Alanine, Valine, Leucine, Isoleucine and Proline.
• Glycine (Gly, G) is the simplest amino acid (R = H)
• Alanine (Ala, A) has a CH3 side chain
• Valine (Val, V) features a branched side chain with an HC(CH3)2 group
• Leucine (Leu, L) exhibits a larger branched side chain with an H3C-CH-CH2- group.
• Isoleucine (Ile, I) is similar to leucine but with a different arrangement, having a CH3-CH2-CH(CH3)- group.
• Proline (Pro, P) has a cyclic structure where the side chain connects back to the amino group
• Alkyl side chains lack functional groups and are hydrophobic

Aromatic Amino Acids

• Aromatic amino acids include Phenylalanine, Tyrosine and Tryptophan.
• Phenylalanine (Phe, F) contains a benzene ring attached to the amino acid backbone.
• Tyrosine (Tyr, Y) is similar to phenylalanine but includes a hydroxyl (OH) group on the benzene ring
• Tryptophan (Trp, W) has an indole ring structure fused to a benzene ring
• Aromatic amino acids are hydrophobic.

Alcohol Containing Amino Acids

• Alcohol-containing amino acids include Serine and Threonine
• Serine (Ser, S) has a hydroxyl (OH) group attached to an aliphatic side chain (CH2OH).
• Threonine (Thr, T) includes a hydroxyl group and an additional methyl group (CH3) on its aliphatic side chain
• Serine and Threonine have aliphatic side chains containing a hydroxyl group, making them hydrophilic (water-loving).
• These can interact with water and other polar groups via hydrogen bonding.
• They contain an alcohol function group and are chemically reactive.

Sulfur Containing Amino Acids

• Sulfur-containing amino acids are Cysteine and Methionine
• Cysteine (Cys, C) contains a thiol (-SH) group
  • The sulfur-containing amino acids are hydrophobic
  • The -SH (thiol) group in cysteine is chemically reactive
• Methionine (Met, M) contains a thioether group (S-CH3)

Acidic Amino Acids

• Acidic amino acids include Aspartic Acid and Glutamic Acid
• Aspartic Acid (Asp, D) has a carboxyl group (COOH) on its side chain
• Glutamic Acid (Glu, E) has an additional methylene group (CH2) before the carboxyl group (COOH)
• Asp and Glu are acidic and very hydrophilic
• They have chemical reactivity and are often found in the active sites of enzymes.

Basic Amino Acids

• Basic amino acids include Lysine, Arginine and Histidine
• Lysine (Lys, K), Arginine (Arg, R) and Histidine (His, H) include nitrogen in their side chains
• Lys, Arg and His are basic, have polar sides, and are hydrophilic
• They are often found in the active sites of enzymes

Amide Containing Amino Acids

• Amide-containing amino acids include Asparagine and Glutamine
• Asparagine (Asn, N) and Glutamine (Gln, Q) contain an amide group (CONH2) in their side chains
• Amides are chemically less reactive than carboxylic acids, but are still polar and therefore hydrophilic.

Essential and Non-Essential Amino Acids

• Essential amino acids must be obtained from diet because plants or microorganisims biosynthesise them.
• Essential amino acids include Arginine (Arg), Histidine (His), Isoleucine (Ile), Leucine (Leu), Lysine (Lys), Methionine (Met), Phenylalanine (Phe), Threonine (Thr), Tryptophan (Trp) and Valine (Val).
• Non-essential amino acids can be biosynthesized within the body (if required) but they are mostly obtained from diet.
• Non-essential amino acids are Alanine (Ala), Asparagine (Asn), Aspartic Acid (Asp), Cysteine (Cys), Glutamic Acid (Glu), Glutamine (Gln), Glycine (Gly), Proline (Pro), Serine (Ser) and Tyrosine (Tyr)

Other Amino Acids

• Additional but rare amino acids exist but are not coded by DNA (thus biosynthesized).
• Examples are 3-Methyl histidine, y-Carboxyglutamate, Dopamine, Histamine
• Amino acids are also used as starting materials for the biosynthesis of important molecules within the body.
• The neurotransmitters Dopamine and Histamine involved in shock and allergic response, are derived from amino acids.

Acidity/Basicity of Amino Acids

• The human body is aqueous (approx. 80% water and buffered at pH 7.4.
• Under these conditions, free amino acids are dipolar and exist as Zwitterions.
• The ionisation state of an amino acid in solution varies with pH
• Side chain functional groups are affected by local pH conditions
  • Acidic side chains are deprotonated at physiological pH
  • Basic side chains are protonated at physiological pH

Stereochemistry of Amino Acids

• Any carbon atom with 4 different substituents is asymmetric or chiral
• Molecules containing a chiral center can rotate plane-polarized light in (+) or (-) directions, thus are optically active.
• A molecule with a chiral carbon can exist as enantiomers
  • These have the same molecular formula but differ in their configuration in space.
• L-amino acids have the same configuration at their chiral carbon as L-glyceraldehyde
• All naturally occurring amino acids have the L-configuration, except glycine
• Some organisms synthesize D-amino acids
• The D and L system is arbitrary and shouldn't be confused with dexotrorotatory (+) and levororotatory (-).
  • L-amino acids can rotate plane polarized light in either directions
  • For example, L-proline (-86.2°) and L-arginine (+12.5°)

Protein Synthesis: Nature

• The central dogma is DNA ⇒ RNA ⇒ Proteins.

Protein Synthesis: Laboratory

• The condensation reaction between the amino and carboxyl group of two amino acids results in the formation of a peptide bond
• A dipeptide [Ala-Ser] means small protein, less than 50 amino acids
• The reaction can be repeated to synthesize a tripeptide (e.g. Ala-Ser-Phe)
• Convention dictates peptides be named/drawn from the amino (N) terminal on the left to the carboxy (C) terminal on the right: e.g H2N-Ala-Ser-Phe-CO₂H

Chemical Steps in Laboratory Peptide Synthesis

• Protection involves blocking the carboxyl and amino groups not involved in the required peptide bond
• Activation involves the carboxyl group to form a peptide bond
• Hydroxide is a poor leaving group
• Activation can be achieved by carboxyl group conversion to an acid chloride, where Chloride (Cl-) is a better leaving group
• Coupling: the free amino group of the second amino acid reacts with the activated carboxyl group to form a new peptide bond
• Deprotection involves removing the protecting groups.

Uses of Peptides

• Aspartame is a synthetic dipeptide 200x sweeter than sugar
• B-Endorphin is an endogenous 31 amino acid neuropeptide with analgesic effects
• Eptifibatide (Integrilin) is a cardiovascular antiplatelet drug
• Some cyclic, pseudo-peptides may contain amino acids and a non-peptide region
• Ciclosporin: is a widely used immunosuppressant, isolated from natural sources. It prevents rejection following organ and tissue transplantation It is a cyclic peptide aht contains naturally occurring and modified (microorganism biosynthesis) amino acids.
• Peptides: can be very useful therapeutic molecules but are also very readily metabolised (broken down) particularly in the stomach.

Explore the crucial link between amino acid side chains and protein structure. Delve into the functions of proteins and the impact of mutations on their structure. Understand the role of amino acids and polypeptide chains in protein formation.