Proteins and Enzymes 1

Proteins and Enzymes 1

Proteins and Enzymes 1

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What are Proteins?

  • Discovered in 1838
  • These molecules were very abundant in biological systems; they were clearly very important, but their precise function was unknown
  • Subsequently determined that proteins were polymers. Nature uses 20 distinct monomers (amino acids) to build all proteins
  • Amino acids fall into seven distinct classes based on the chemical properties of the side chain (R): aliphatic, aromatic, alcoholic, sulfur-containing, acidic, basic or amide
  • The amino acid side chain provides a specific chemical property to the protein
  • Each amino acid is identified by a three letter code or a one letter code
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What are Proteins?

  • Bonding between carboxylic acid and amino groups of two adjacent amino acids forms a continuous peptide backbone with protruding R groups
  • The continuous peptide backbone folds into a specific three dimensional shape
  • Every protein has a unique and complex structure
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Biological Functions of Proteins

  • The complex structure of proteins allows them to undertake a wide variety of functions
    • Catalysis: Proteins that catalyse chemical reactions in the body are known as enzymes - several thousand known
    • Storage and transport: Ferritin stores and transports iron in the body. Haemoglobin transports oxygen
    • Mechanical support and shape: Collagen is a component of supportive tissue, e.g. ligament, skin, bone etc.. Most abundant protein found in vertebrates
    • Decoding information, gene expression: RNA polymerase synthesises RNA (directed by DNA)
    • Specialist functions: immunoglobins (antibodies) and some hormones (e.g. insulin) are proteins
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Amino Acids

  • Aliphatic amino acids contain alkyl sidechains
    • The aliphatic amino acids include the simplest amino acid, glycine, R = H
    • The alkyl side chains contain no functional groups, but are very hydrophobic
  • Aromatic amino acids contain an aromatic ring
    • The aromatic amino acids Phe, Trp and Tyr are quite hydrophobic
  • Alcohol containing amino acids
    • Ser and Thr have aliphatic side chains containing a hydroxyl group. As a result they are hydrophilic since they can interact via hydrogen bonding with water and other polar groups. They contain a functional group (alcohol) and are chemically reactive
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Amino Acids

  • The sulfur containing amino acids are hydrophobic in character
    • The -SH (thiol) group in cysteine is very chemically reactive
  • Acidic amino acids
    • Asp and Glu are acidic and very hydrophilic. Also, they have chemical reactivity and are often found in the active sites of enzymes
  • Amide containing amino acids
    • Amides are neutral and chemically less reactive than carboxyl acids. They are still very polar and therefore hydrophilic
  • Basic (nitrogen containing) amino acids
    • Lys, Arg and His are basic; they have polar sides so they are very hydrophilic. They are often found in the active sites of enzymes
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Essential Amino Acids

  • Essential amino acids are only biosynthesised by plants or microorganisms. These must be obtained from the diet
    • Arginine (Arg)
    • Histidine (His)
    • Isoleucine (Ile)
    • Leucine (Leu)
    • Lysine (Lys)
    • Methionine (Met)
    • Phenylalanine (Phe)
    • Threonine (Thr)
    • Tryptophan (Trp)
    • Valine (Val)
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Non-Essential Amino Acids

  • Non-essential amino acids can be biosynthesised within the body if required but are mostly obtained from the diet
    • Alanine (Ala)
    • Asparagine (Asn)
    • Aspartic acid (Asp)
    • Cysteine (Cys)
    • Glutamic acid (Glu)
    • Glutamine (Gln)
    • Glycine (Gly)
    • Proline (Pro)
    • Serine (Ser)
    • Tyrosine (Tyr)
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Other Amino Acids

  • Other amino acids, in addition to the the common 20 do exist. However, they are generally very rare and are not coded by DNA (they are biosynthesised)
  • 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
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Acidity/Basicity of Amino Acids

  • The human body is mostly aqueous (approximately 80% water) and buffered at pH 7.4 (physiological pH)
  • Under these conditions, free amino acids are dipolar and exist as 'Zwitterions'
  • For an amino acid in solution the ionisation varies with pH
  • The side chain functional groups are also affected by local pH conditions
  • At physiological pH, acidic side chains are deprotonated and basic side chains are protonated
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Stereochemistry of Amino Acids

  • Any carbon atom with 4 different substituents is asymmetric or chiral
  • Molecules containing a chiral centre are able to rotate plane polarised light in a clockwise (+) or anti-clockwise (-) direction: they are optically active
  • A molecule with a chiral carbon can exist as enantiomenrs: compounds with the same molecular formula but differing in their configuration in space
  • To describe different enantiomers, pairs of amino acids are designated D or L. The reference compound is glyceraldehyde (a sugar)
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Stereochemistry of Amino Acids

  • L-amino acids have the same configuration at their chiral carbon as L-glyceraldehyde
  • All the naturally occurring amino acids have the L-configuration, except glycine. Some organisms synthesise D-amino acids

Protein Synthesis: Laboratory

  • A condensation reaction between the amino group and carboxyl group of two amino acids results in formation of a peptide bond
  • A dipeptide is formed; a peptide is a small protein of less than 50 amino acids
  • This reaction can be repeated to synthesise a tripeptide
  • Peptides should be named/drawn from the amino (N) terminal on the left to the carboxy (C) terminal on the right

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Chemical Steps in Laboratory Peptide Synthesis

  • Protection: Blocking of the carboxyl and amino groups not involved in the required peptide bond
  • Activation of the carboxyl group involved in peptide bond formation
    • The carboxyl group can be activated by conversion to an acid chloride
  • Coupling: Reaction of the free amino group of the second amino acid with the activated carboxyl group to form a new peptide bond
  • Deprotection: Removal of the protecting groups
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Chemical Steps in Laboratory Peptide Synthesis

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Chemical Steps in Laboratory Peptide Synthesis

Uses of Peptides

  • Aspartame: synthetic dipeptide 200x sweeter than sugar
  • B-endorphin: an endogenous 31 amino acid neuropeptide with analgesic effects
  • Eptifibatide (Integrilin): cardiovascular antiplatelet drug  is a cyclic pseudo-peptide (i.e. contains amino acids and a non-peptide region
  • Ciclosporin is a widely used immunosuppressant drug, isolated from natural sources, that prevents rejection following organ and tissue transplantation. It is a cyclic peptide that 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
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Chemical Bonding

  • Ionic bond: complete charge separation, e.g. sodium chloride, no sharing of electrons
  • Covalent bond: sharing of electrons, organic compounds e.g. phenylalanine
  • Hydrogen bonds: weak bonds that form between a hydrogen atom attached to an electronegative atom and another electronegative element, e.g. oxygen or nitrogen
  • Hydrogen bonds are weak (1-5% of a covalent bond) but are vitally important to biological systems. Hydrogen bonding in water as responsible for its liquid state
  • Hydrogen bonds hold proteins and DNA together
  • Many drugs interact with their targets by hydrogen bonds
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Hydrophobic Interactions and Bonding in Proteins

  • Hydrophobic interactions are the attraction observed between electrically neutral molecules. Electron movements in covalent bonds causes small dipoles to develop
  • Covalent: Most common in the backbone, occassionally seen between two sidechains - strong
  • Ionic: Attraction between charged groups, also known as a 'salt bridge' - moderate
  • Hydrogen bond: Very important for protein structure - weak
  • Hydrophobic: Exclusion of water, a driving force of this interaction - very weak
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