Cell Components Revision

Table of Contents

CELL COMPONENTS REVISION....................................................................................................................... 1

NUCLEIC ACIDS AND PROTEINS...............................................................................................................................1
AMINO ACIDS....................................................................................................................................................10
PROTEINS......................................................................................................................................................... 18
ENZYMES..........................................................................................................................................................24
VITAMINS.........................................................................................................................................................32
ENERGY AND METABOLISM..................................................................................................................................43

Nucleic Acids and Proteins

Nucleosides and Nucleotides
- DNA and RNA are molecules that play a fundamental role in the
storage of genetic info
- Nucleic acids are polymers that are made up of nucleotides
- Nucleotides can structurally be divided into 3 parts: heterocyclic
base, a sugar, and phosphate
- The most notable difference between DNA and RNA nucleotides is
the sugar: deoxyribose in DNA and ribose in RNA
- Nucleoside is a nucleotide lacking a phosphate group
Purines and Pyrimidines
- The bases in nucleosides and nucleotides are either monocyclic
pyrimidines or bicyclic purines
- The pyrimidine bases are
cytosine, thymine, and uracil
- The purine bases are adenine
and guanine
- Thymine is found only in DNA
- Uracil is found only in RNA

Sugar Unit in Nucleosides

- The sugars in DNA and RNA are pentoses
- They are D-ribose in RNA and 2-deoxy-D-ribose in DNA
- In all cases, the sugar is present in a five-membered acetal ring form

Glycosidic Bond
- The nucleobase is linked to the sugar unit through an N-glycoside
bond at C1
- The linkage is always β
- Purines are linked through N9 while
pyrimidines are linked through N1
- Primed numbers are used for the sugar
and non-primed numbers are used for the nucleobase atoms

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DNA and RNA Nucleosides

Phosphate Group in Nucleotides

- The phosphate group in nucleotides is attached via a phosphor
ester linkage
- It may be attached to the 5’- or 3’-

DNA
- DNA is made up of long unbranched chains of
oligonucleotides which are linked via a phosphate
group that joins the sugar unit with the nucleobase
- The ester linkage between nucleotides is often a
phosphodiester bond
- The phosphodiester bond links a 5’- of one nucleotide
to the 3’ of the next nucleotide
- Thus, nucleic acids have two ends: 5’- and 3’- ends

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 - In some organisms (some bacteria and viruses) the 5’- and 3’- ends
of DNA are linked to give circular DNA
- The base sequence of DNA is written from the 5’- to the 3’- end
(e.g. 5’-ATCCGATGG-3’)
- The nucleobases in DNA have the ability to form H bonds
between themselves
- This property is essential to the double-helix
arrangement of DNA, translation, and transcription via
RNA
- The poly nucleotide chain of DNA coils into a helix which
gets bonded to another helical strand by H bonds
between the appropriate base pairs
- In DNA, the pairs are A-T and C-G
- A-T are bonded by 2 H bonds while C-G are bonded by 3 H bonds

- The DNA double helix has 2 poly nucleotide chains twisting on a

common axis
- The bases are directed inwards to allow H bonding
(base pairing)
- The sugar units and the phosphodiester bonds off
the main chains will form the outside part of the
double helix
- The planes of the base pairs are perpendicular to
the helix axis
- The helix makes a complete turn every 10 bases
- The 2 chains are complementary in sequence – if you know the
identity of on chain, you can work out the second chain

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 - The chains are antiparallel
- Since the glycoside bonds between the sugars and bases of a
particular base pair are not directly opposite to each other, grooves
along the outside of the double helix array are unequal in with
giving rise to what is known as the minor groove and major groove
- Grooves mainly contain water molecules, but they are
distinguishable to (anticancer) small molecules

DNA Replication
- In cell division, the DNA molecule is replicated so that each
daughter cell will carry its own DNA molecule
- DNA replication proceeds in 3 enzymatically mediated steps:
initiation, elongation, termination

- Enzymes involved in DNA replication

o Ligases
o Primase
o DNA polymerase
o Helicase
o Exonuclease

- Initiation
o Initiated at points within the DNA strand known as origins
o Multiple origin sites exist within the DNA structure; when
replication begins these sites are called replication forks
o DNA helicase unwinds the double helix and exposes each
strand so they can be used as a template for replication
o It does this by hydrolysing the ATP used to form the bonds
between the nucleobases
o DNA primase synthesises a small RNA primer which acts as a
kick starter for DNA polymerase
- Elongation
o Once DNA polymerase is attached to the template strands it’s
able to start synthesising new strands of DNA

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 o DNA polymerase is only able to extend the primer by adding
free nucleotides to the 3’ end
o The newly formed strand is called the leading strand
o The other template strand (lagging strand) is antiparallel and
is read 5’ to 3’ but DNA polymerase cannot add bases to the
5’ end
o Instead, as the helix unwinds, RNA primers are added to the
newly exposed bases and DNA synthesis occurs in fragments
(Okazaki fragments)
- Termination
o The process continues until there is no more DNA strand left
(end of the chromosome) or until 2 replication forks meet and
subsequently terminate
o Once replication has finished, the newly synthesised strands
are bound and stabilised
o For the lagging strand, RNAase H removes the RNA primer at
the beginning of each Okazaki fragment and DNA ligase joins
these fragments together to create 1 complete strand

- The precursors for the synthesis of new DNA strands are nucleoside
triphosphates
- These triphosphate anhydrides are susceptible to nucleophilic
attack from hydroxy groups

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 - DNA chain extension is simply an esterification reaction of the 3’-
hydroxyls using the triphosphate anhydrides that have the
diphosphate as a good leaving group
- The correct triphosphate is selected because of the H bonding
properties of the base pairs

RNA
- RNA differs structurally from DNA in 3 important ways:
o Sugar is ribose
o Uracil not thymine
o Usually single stranded
- DNA stores genetic info and RNA participates in the processes by
which this info is used
- 3 major forms in prokaryotic cells: mRNA, tRNA, rRNA

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The Genetic Code
- It’s the sequence of bases along one
DNA strand (the coding strand) which
provides info for the synthesis of
proteins in an organism
- A complementary second strand exists,
and this is termed the template strand
- A gene is a DNA segment that contains
the info necessary for the synthesis of
one protein

- Each amino acid in a protein is specified by

a sequence of 3 nucleotides called a codon
- Every amino acid is designated 2 or 3
specific triplet codons
- The signal for starting translation is the
codon of methionine, which has only one
codon
- Three different codons are known as stop
codons, which stop the translation process

Transcription
- Although the amino acid sequence of a protein is defined by the
sequence of codons in DNA,
it is RNA that participates in
the interpretation of this
sequence
- The synthesis of mRNA from
the DNA template is called
transcription
- First, DNA is unwinded and the template strand is the one used for
mRNA synthesis
- ATP, CTP, GTP, UTP are used in building the mRNA sequence
- Process is mediated by RNA polymerase
- Synthesis occurs in the nucleus

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Translation
- After synthesis, mRNA moves from the nucleus to
the cytoplasm and then the ribosome
- Ribosomes are made up of 2 subunits termed
(60S/40S + 50S/30S)
- which are combinations of rRNA and proteins
- A tRNA is specific for a particular amino acid
- One arm on tRNAs always has CCA to which
amino acids are ligated

- The mRNA sequence is

read from the 5’ to the 3’ in translation
- The codon and anticodon are bonded by H bonds
- In prokaryotes, the first amino acid
encoded is N-formylmethionine, which
has the same codon as methionine
(AUG)
- Note that the peptide is synthesised
from the N-terminal to the C-terminal

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 - The initiator aminoacyl-tRNA is bound and positioned in the P site
- The next amino acyl-tRNA is bound and positioned in the adjacent A
site
- Peptide bond is formed
between amino acid in A site
and that in P site
- The peptide chain is now
attached to the tRNA in site A
- The empty tRNA in site P is
release from the ribosome
- The peptide-tRNA in site A now moves to site P and a new
aminoacyl-tRNA moves into site A, etc.

Amino Acids
- Abundant in biological systems
- Proteins are polymers
- 20 distinct monomers (amino acid) to build all proteins
- Amino acids fall into 7 distinct classes based on the chemical
properties of the sidechain (R): aliphatic, aromatic, alcoholic, sulfur
containing, acidic, basic, amide
- The amino acid sidechain provides a specific chemical property to
the protein
- Each amino acid is identified by the 3-letter code or a one letter
code

- Bonding between carboxylic acid and amino groups of 2 adjacent

amin acids forms a continuous peptide backbone with protruding R
groups

- The continuous peptide backbone folds into a specific 3D shape

- Every protein has a unique and complex structure

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 Biological Functions of Proteins
- Catalysis – proteins that catalyse chemical reactions in the body are
known as enzyme and several thousand are known
- Storage and transport – e.g. ferritin stores and transports iron in
the body and haemoglobin transports oxygen
- Mechanical support and shape – e.g. collagen is a component of
supportive tissue (e.g. ligament, skin, bone, etc.)
- Decoding info, gene expression – RNA polymerase synthesises RNA
(directed by DNA)
- Specialist functions – Immunoglobins (antibodies) and some
hormones (e.g. insulin) are proteins

Aliphatic Amino Acids

- The aliphatic amino acids
include the simplest amino
acid – glycine R = H
- They contain alkyl side
chains with 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

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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 with water
and other polar groups via H bonding
- They are chemically reactive

Sulfur Containing Amino Acids

- Hydrophobic
- The -SH (thiol) group in
cysteine is very chemically
reactive

Acidic Amino Acids

- Asp and Glu are acidic
and very hydrophilic
- Are chemically reactive
- Often found in active sites
of enzymes

Basic (Nitrogen Containing) Amino Acids

- Lys, Arg and His are basic
- Have polar sides so very hydrophilic
- Often found in active
sites of enzymes

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Amide Containing Amino Acids
- Neutral and chemically less reactive than carboxylic acids
- Still very polar and hydrophilic

Essential/Non-Essential Amino Acids

- Some amino acids are only
biosynthesised by plants or
microorganisms
- These must be obtained from the
diet and are known as essential
amino acids
- Non-essential amino acids can be
biosynthesised within the body if required however they are mostly
obtained from the diet

Other Amino Acids

- Other amino acids in addition to the common 20 do exist
- 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 in the body
- Neurotransmitters
dopamine and
histamine are derived
from amino acids

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Acidity/Basicity of Amino Acids
- The human body is mostly aqueous and buffered at pH 7.4
- Under these conditions free amino acids are dipolar and
exist as zwitterions
- For an amino acid in solution the ionisation state 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

Stereochemistry of Amino Acids

- Molecules with a chiral center are able to rotate plane polarised
light in a clockwise (+) or anti-clockwise (-) direction – they are
optically active
- To describe different enantiomers, pairs of amino acids are
designated D or L with the reference compound glyceraldehyde (a
sugar)

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 - 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 in a Laboratory

- A condensation reaction between the amino group and carboxyl
group of 2 amino acids results in formation of a peptide bond

- A dipeptide (Ala-Ser) has been formed

- A peptide is a small protein of less than 50 amino acids
- This reaction can be repeated to synthesise a tripeptide

- By convention, peptides should be named/drawn from the amino

(N) terminal on the left to the carboxy (C) terminal of the right (e.g.
H2N-Ala-Ser-Phe-CO2H)

Chemical Steps in Lab Peptide Synthesis

- Protection – blocking of the carboxyl
and amino groups not involved in the
required peptide bond
- Activation – activating the carboxyl
group involved in peptide bond formation

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 - 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 protecting groups

Uses of Peptides

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 - 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, particularly in the stomach

Proteins
- Proteins are large molecules composed of several hundred amino
acids
- The linear sequence of amino acids in a peptide is referred to as the
primary structure
- A polypeptide chain is not linear and folds into a biologically active
shape
- The biologically active form is known as the native conformation
- The biological functions of many proteins can be explained on the
basis of their conformations or shapes

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 - E.g. an enzyme fold to form an active site that can recognise
substrate molecules

Factors Effecting Protein Conformation – The Peptide Bond

- The properties of the peptides bond have considerable impact on
the shape and function of proteins
- The peptide bond is planar, electron resonance gives 40% double
bond character
- The peptide bond may be regarded as the average of 2 extreme
resonance forms

- Some properties of the peptide bond are a result of its double bond

character
- It is rigid and planar
- Rotation around the peptide bond is not possible
- Peptide bonds have trans conformation
- Steric hindrance between side chain
groups favours the trans conformation
- Since the peptide bond is rigid, only 2
free movements exist in a polypeptide
chain:
o Rotation around the αC-N bond is
called the phi () torsion angle
o Rotation about the αC-C bond is
called the psi () torsion angle

 and  Bonds – Restricted Conformation

- Protein conformation depends on phi and psi rotation
- The flexibility of these bonds allows the primary sequence to fold
into its native conformation
- Rotation is limited by:

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 o Steric hindrance: bulky groups e.g. side chains cannot
approach each other
o Rigidity of the peptide bond restricts movement
o Favourable interactions (e.g. H bonds) with other regions of
the polypeptide chain

Secondary Structure – The α Helix

- The 3D dimensional arrangement of primary amino acid
sequence is called the secondary structure of a protein
- The α helix is a secondary structure that results when
consecutive amino acid residues have similar phi and psi
torsion angle values
- Phi = --57˚, Psi = --47˚
- The α helix is a single helix not to be confused with DNA
- 3.6 amino acid residues are required for one complete
turn of the helix
- Each backbone carbonyl oxygen is H bonded to the
peptide nitrogen of the 4th residue along (towards C
terminus)
- This is a favourable interaction that stabilises the helix
- Although H bonds are weak, they hold the helix
structure together
- Side chains are arranged on the outside of the helix

- Receptors are proteins rich in α helices

- They usually contain a trans-membrane domain composed entirely
of α helices

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 - Crystal structure of the adenosine receptor (trans membrane
domain) prevalent in cardiac tissue:

- Adenosine, a treatment for supraventricular tachycardia, binds to

the trans membrane domain

Secondary Structure - β Sheet

- A β sheet is a secondary protein structure in which several strands
(called β strands) of the peptide backbone are H bonded to
themselves
- The β sheet is an elongated reasonably flat ‘sheet-
like’ structure
- Inter-strand H bonds between backbone carbonyl
oxygen and amide nitrogens stabilise the sheet
- In the β sheet side chain interactions (mostly
hydrophobic interactions between small groups)
can provide additional stabilisation
- They come in 2 varieties:
o Antiparallel β sheet – optimally H bonded,
linear H bonds (better overlap = stronger
bond), 2-15 strands possible (average 6)
o Parallel β sheet – H bonds distorted, sheet less
stable, no more 5 strands encountered

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Fibrous Proteins
- Contain only α helix secondary structure
- Simple, elongated structure resembling threads or fibres
- Provide mechanical support in skin, tendons, bones
- Physically durable, chemically inert and water insoluble
- Structure maintained by H bonding within the α helix

- Hair is composed mostly of α keratin, a double coil of α helices

- Many double coils are packed together to form a strand of hair

Tertiary Protein Structure

- Refers to the 3D arrangement of secondary structure
- Tertiary protein (e.g. enzymes) usually contain an
assortment of secondary features
- Receptors also have a tertiary structure
- The trans membrane domain is usually attached to a
cytoplasmic domain (i.e. a mixture of secondary structure)
- The precise structure of membrane bound receptors is
extremely difficult to determine

Globular Proteins
- Tertiary proteins with greater structural diversity than fibrous
proteins
- All enzymes are globular proteins
- Some properties of globular proteins

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 o Water soluble
o Compact, roughly spherical
o Tightly folded peptide chains
o Hydrophobic interior, hydrophilic surface
o Structure maintained by covalent and H bonding, non-
covalent crosslinks, and hydrophobic interactions
o Possess indents or clefts (active site)
o Enzymes

Stabilisation of Tertiary Structure – Hydrophobic Effect

- Proteins are more stable in water with their hydrophobic side
chains tucked into the protein interior
- Non-polar substances will always minimise their contact with water
- Non-polar side chains aggregate causing the protein to fold with
non-polar side chains inside and polar side chains outside the
protein in contact with water
- Efficient packing maximises van der Waals interactions between
non-polar residues and excludes water from the interior of the
protein
- Structure control’s function: enzymes must be water soluble to
function in cells
- Val, Leu, Ile, Met, Phe and Ala are rarely encountered on protein
exteriors

- Proteins
are arranged so virtually all possible H bonds are formed

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 - Polar side chains forced into the protein interior can neutralise their
polarity by forming H bonds of electrostatic interactions

Stabilisation of Tertiary Structure – Covalent Interactions

- Disulfide bonds are covalent cross links that form between adjacent
cysteine residues and help stabilise the conformation of some
proteins
- A covalent bond is very strong compared to H bonds and van der
Waals
- Disulfide links are especially common in proteins that are secreted
from cells

Effects of Temp and pH on Tertiary Structure

- The tertiary structure of an enzyme is responsible for biological
activity
- Tertiary structure is maintained by weak interactions (mostly H
bonds and vdw)
- Variations of pH and temp disrupt the stabilising interactions
causing changes to the tertiary structure
- The protein is said to be denatured

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 - Enzymes have evolved to function (i.e. max. stabilisation of tertiary
structure) at physiological conditions pH 7.4 and 37˚C

Quaternary Structure
- Refers to proteins that are composed of more
than one polypeptide strand
- Haemoglobin is composed of 4 globular
protein subunits: 2 identical α units
containing 141 amino acids and 2 identical β
units containing 146 amino acids
- Each subunit contains an iron atom vital for
the transport of oxygen in the blood
- Insulin consists of 2 peptide chains linked and maintained in the
biological active conformation by 3 disulfide bridges

Enzymes
- Catalysts for biochemical reactions
- Virtually every chemical reaction in a biological system is regulated
by an enzyme
- Catalysts do not change the outcome of a reaction but effects the
rate of a reaction
- Catalysts are chemically unaltered by the reactions
- Enzymes accelerate the rate of a biochemical reaction by lowering
the activation energy for that reaction
- Activation energy of enzyme catalysed reactions is lower than that
of an uncatalysed reaction

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What are Enzymes?
- Globular proteins (tertiary structure)
- Tertiary structure creates active site pockets – reactants fit tightly
and interact with the protein (lowering activation energy)
- Globular proteins can bind one or more substances in an active site
– the substrate(s) and any cofactors that may be required for the
reaction

Enzyme Nomenclature
- Named and classified according to the reactions catalysed
- The suffix -ase is added to the name of the substrate or a
descriptive term for the type of reaction
- The International Union of Biochemistry catergorises enzymes into
6 major groups according to the general class of chemical reactions
they catalyse:
o Oxidoreductases
o Transferases
o Hydrolases
o Lyases
o Isomerases
o Ligases
- The Enzyme Commission assigns a unique code number to each
enzyme:

Enzymic Energy Profile

- Enzymes lower the activation
energy (the barrier to the reaction)

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 - Physically, this is achieved by the correct and precise orientation of
the substrate within the enzyme active site

Properties of Enzymic Reactions

- Fast – most reactions are at least a million times faster
- Specific for the substrate – enzymes can distinguish between
functional groups, isomers, and enantiomers
- Efficient – enzymes have evolved to minimise ‘waste’ by-products

Enzyme Cofactors
- Enzymes are composed of 20 amino acids
- There is limited chemistry possible with 20 different side chains
- Cofactors supply chemical groups not otherwise found in active
sites
- Many enzymes are inactive as a protein alone (apoenzymes) and
cofactors are required for activity
- Coenzyme – a non-protein organic compound, produced in living
cells, which is involved in the activation of enzymes
- Cosubstrates – weakly bound to enzyme, altered during the course
of the reaction, and dissociate from the active site, regenerated in
another enzymic reaction and recycles
- E.g. ATP  ADP, NAD+  NADH
- Prosthetic groups – tightly bound to the
enzyme but must be regenerated each
catalytic cycle
- E.g. thiamine pyrophosphate (Vit B1)
and haem

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Coenzyme Reaction

-
-
-
-
Unlike enzymes, the coenzyme is often structurally altered during an
enzyme-catalysed reaction
- In the top reaction, the functional group X is cleaved from
molecules A and bound to the coenzyme producing the active form
of the coenzyme
- Coenzyme-X then acts as a coenzyme for the second enzyme
allowing the functional group X to be transferred onto molecule B
- In the body, the coenzyme will be regularly regenerated by the first
enzyme so that it can act as a coenzyme for the second enzyme
multiple times

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Essential Ions
- Nearly 1/3 of proteins require a metal ion for activity
- They are known as metalloproteins or metalloenzymes
- Fe2+ found in haem (haemoglobin, myoglobin)
- Mg2+ found in many kinases (reactions involving phosphorylations)
- Zn2+ found in many enzymes (oxidations, reductions, DNA
recognition “zinc fingers”)
- Copper, manganese, cobalt, molybdenum are other trace metals
used by enzymes
- Recommended daily intake of cobalt is 2µg (as vit B12) – deficiency
leads to pernicious anemia (low RBC, neurological deterioration)

Oxidoreductases
- Catalyse oxidation and reduction (redox) reactions
- Include dehydrogenases, oxidases, peroxidases, reductases
- Alcohol dehydrogenase [1.1.1.1] – oxidises ethanol to ethanal using
cofactor, NAD+ and zinc
- NAD+ is the biologically active form of vit B3 (niacin)

- Alcohol dehydrogenase mechanism

- The enzyme orientates all species correctly for the reaction to occur

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Transferases
- Catalyse functional group transfer reaction
- Includes kinases – important family of enzymes that catalyse
phosphorylation reactions
- Transcarboxylase [2.1.3.1] catalyses
the simultaneous conversion of
propionyl CoA to methyl malonyl CoA
(top) and oxaloacetate to pyruvate
(bottom)
- Uses biotin (vit H) cofactor
- Biotin is a cofactor frequently involved in enzymic carboxylation
reactions because of its ability to bind and manipulate a carboxylate
group

Hydrolases
- Catalyse hydrolysis reactions
- Trypsin [3.4.21.4] mediates the hydrolysis of Lys-aa or Arg-aa
peptide bonds
- Trypsin is a serine protease enzyme – it needs no cofactor, its active
site contains sufficient functionality to be able to perform the
reaction unassisted (Asp-His-Ser: a catalytic triad)

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 - Trypsin mechanism of action:

1. The serine residue attacks the substrate peptide carbonyl bond,

aided by Asp and His which promote the nucleophilic attack and
stabilise the resulting positive charge
2. The tetrahedral intermediate collapses…
3. Liberating R-Nh2, but the carboxyl residue remains bonded to the
enzyme via the serine (an ester linkage, easily hydrolysed)
4. A water molecule, correctly orientated by interaction with His,
attacks the ester carbonyl group
5. Reaction complete – active site amino acids are unchanged and
ready for further catalysis, new amino and carboxylic groups
exposed by hydrolysis

Lyases
- Catalyses bond breaking reaction (but not hydrolytic or oxidative
bond breaking)
- L-Dopa decarboxylase [4.1.1.28] – decarboxylation of L-amino acids
- Coenzyme involved – pyridoxal phosphate (vit B6, R = CH2OPO32-)

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 - Complex mechanism (abbreviated below) – the aldehyde group
reacts with amines (drug) to form an imine (-CH=N-), aldehydes not
available in amino acid side chains therefore vitamin required

Isomerases
- Catalyse isomerisation reactions (i.e. rearrangement of groups
within a substrate molecule)
- Alanine racemase [5.1.1.1] found in bacteria – catalyses the
interconversion of L and D alanine
- D alanine essential for bacterial cell wall
- Coenzyme involved – pyridoxal phosphate (vit B6)

- Enzyme removes H from one face of amino acid (A) and replaces on
opposite face (B)

Ligases
- Catalyse the ligation (joining) of 2 substrates
- Pyruvate carboxylase [EC 6.4.1.1] joins pyruvate with carbon
dioxide to generate oxaloacetate – this is an important step in
metabolism/the Krebs cycle

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Vitamins
Macro and Micronutrients
- Macro – carbs, fats, proteins
- Micro – minerals and vitamins
- Minerals – calcium, magnesium, sodium

Vitamins
- General term for any of several organic substances essential for
normal metabolic processes and which, when absent in the diet,
produce deficiency states as they aren’t produced naturally by the
body
- Many coenzymes are synthesised from dietary precursors (which
are often vitamins)
- Required for growth, reproduction, and normal body function
- The term vitamin was first used in 1912 to describe a ‘vital amine’
derived from rice husks that cured beriberi
- Further discoveries were also called vitamins (although not all
amines)
- Excessive intake of some vitamins can cause toxicity
- Classes of vitamins:
o Water soluble
 Readily excreted via kidney in urine
 Required daily in small amounts
 E.g. vits B and C

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 o Lipid soluble
 Stored in the body (usually liver)
 Excessive intake can cause toxicity
 E.g. vits A, D, E and K

Water Soluble Vitamins

- B Vitamins
o Originally thought to be a single vitamin but now understood
to be multiple vitamins (all of which form coenzymes)
o Named in order of discovery but some were later found to be
produced by the body so are no longer classed as vitamins
(e.g. B4, B8, B10, B11)
o Classed together as they’re found in similar foods though
differences in structure and function exist
o Some can be synthesised within the body by intestinal flora
(not the same as being produced BY the body)

Vitamin B1 (aka thiamine)

- Found in cells, therefore present in all whole natural foods (e.g.
whole cereal grains, beans, fruits, yeast
- Coenzyme:
o Thiamine pyrophosphate (TPP)
o Coenzyme to several enzymes in
carbohydrate metabolism pathways
(helping to release energy from food)
o Also involved in conduction of action potentials in neurons
and neuro-muscular transmission
- Deficiency state:
o Prevalent in eastern Asia and in chronic alcoholics
o Athletes and pregnant/lactating women may require
supplements due to increased carbohydrate consumption
o Beriberi can affect different organ systems
o Wet beriberi affects the cardiovascular system and presents
with cardiac failure, dyspnoea and oedema

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 o Dry beriberi affects the peripheral nervous system and
presents with peripheral neuritis, paralysis, and wastage
o A severe form of dry beriberi is Wernicke-Korsakoff syndrome
and is characterised by paralysis of eye movement, unusual
movements, and impaired mental function

Vitamin B2 (aka riboflavin)

- Found in dairy products, eggs, green veg, almonds
- Coenzymes:
o Flavin mononucleotide (FMN)
o Flavin adenine dinucleotide (FAD)
o Heterocyclic system acts as a H acceptor
(or donor)
o Coenzymes act as prosthetic groups on a
family of mainly oxidoreductase enzymes
known as flavoproteins
o Involved in metabolism of fats, carbs, and
proteins
o Also acts as coenzyme in processing other
vitamins (e.g. conversion of vit B6 into its coenzyme
- Deficiency state:
o Rare in developed countries due to fortified foods but
common in developing countries due to malnutrition
o Symptoms include inflammation of mouth and lips
(stomatitis) which is similar to pellagra but without the
widespread skin lesions (pellagra sine pellagra)
o It can also reduce iron absorption leading to anaemia with the
size and haemoglobin content of RBCs remaining normal
(normochromic normocytic anaemia)

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Vitamin B3 (aka nicotinamide – niacinamide, niacin – nicotinic acid)
- Found in meat, fish, nuts
- Nicotinamide also used in acne treatments
- Coenzymes:
o Nicotinamide adenine dinucleotide
(NAD)
o Nicotinamide adenine dinucleotide phosphate (NADP)
o Important in oxidoreductase
enzyme reactions
o NAD(P)+ is an oxidising agent
whilst NAD(P)H is a reducing
agent
o NADPH is important in
anabolic processes (e.g. lipid
and nucleic acid synthesis)
o NAD+ is vital to catabolic
processes (e.g. metabolism of energy sources in fatty acids
and glucose)
- Deficiency state:
o Body not entirely dependent on dietary intake as nicotinic
acid can be produced from dietary tryptophan in vivo
o Deficiency common in areas in which maize is the principal
foodstuff as it is low in both nicotinic acid and tryptophan
o Pellagra – characterised by dermatitis, diarrhoea, dementia
o Chronic alcoholics also at risk of deficiency
- Toxicity state:
o Skin flushes, liver damage
o Mainly seen with niacin supplementation, nicotinamide may
be given as alternative

Vitamin B6 (aka pyridoxine, pyridoxal, pyridoxamine)

- Found in most vegetables (pyridoxine)
or animal origin (pyridoxal,
pyridoxamine)

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 - Interconvertible via their phosphates in vivo
- Coenzyme:
o Pyridoxal-5’-phosphate
o Involved in tryptophan metabolism (vit B6
deficiency is risk factor for vit B3 deficiency)
o Involved in production of neurotransmitters
(e.g. dopamine, GABA)
o Acts as coenzyme for extremely broad range of enzymes
- Deficiency state:
o Rare as most diets contain adequate amounts and some is
synthesised by intestinal flora
o Results in disorders of CNS, skin, mucous membranes
o Patients taking medicines such as isoniazid may benefit from
increased intake due to increased excretion of pyridoxine
- Toxicity state:
o Leads to nerve damage (particularly spinal ganglia) which
manifests as pain/numbness in extremities or in extreme
cases difficulty with motor functions

Vitamin B7 (aka biotin)

- Found in egg, avocado, yeast, fresh veg
- Coenzyme:
o N/A as biotin acts as a coenzyme
o Required by several carboxylase enzymes
that are involved in fatty acid synthesis, amino acid
breakdown and glucose synthesis (gluconeogenesis)
- Deficiency state:
o Severe deficiency never reported in healthy individuals eating
a normal mixed diet
o Presents with thinning hair, brittle nails, rashes, neurological
symptoms
o Pregnant/lactating women at risk of deficiency (clinical reason
unknown)

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 o Those eating lots of raw eggs are at risk due to avidin in the
whites reducing biotin absorption

Vitamin B9 (aka folic acid)

- Found in nuts, seeds, chickpeas, green veg
- Folic acid is the vitamin most commonly
added to ‘fortified’ foods such as flour
- Coenzymes:
o Tetrahydrofolate (THF)
o Coenzyme in enzymatic reactions
that transfer hydroxymethyl
(-CH2OH), formyl (-CHO) and methyl groups in a large no. of
reactions
o Synthesis of amino acids and purine/pyrimidine bases in the
formation of DNA
- Deficiency state:
o Patients with malignant disease or are
pregnant/breastfeeding are at risk of deficiency due to
increased folic acid demand
o A no. of medication (e.g. methotrexate) interfere with folic
acid processing and so create a risk of deficiency
o Folic acid deficiency during pregnancy is associated with low
birth weight, premature birth, and neural tube defects
o Supplements are given in pregnancy, leukaemia, and drug-
induced folate deficiency
o Deficiency can cause megaloblastic anaemia (large RBCs) due
to faulty erythrocyte multiplication and maturation
o Supplementation with folic acid may mask vit B12 deficiency
which leads to damage to the nervous system

Vitamin B12 (aka cobalamin)

- Found in animal sources (meat, fish, milk, eggs)
- Rare in common plant-based food (some in fermented
foods and seaweed)

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 - Coenzyme:
o Adenosylcobalamin
o Methylcobalamin
o Body converts hydroxocobalamin and cyanocobalamin from
the diet into these active forms
o Acts as a cofactor for methionine synthase which is involved
in the production of THF from folic acid
o Also acts as cofactor in molecular rearrangement reactions
(e.g. metabolism of branched-chain amino acids) particularly
in the CNS
- Deficiency state:
o Causes pernicious anaemia – megaloblastic anaemia plus
gastrointestinal (diarrhoea, loss of bladder control) and
neurological symptoms (seizures, degeneration of spinal cord)
o Folic acid supplementation can reverse the megaloblastic
anaemia but not reverse other symptoms
o Care must be taken with anaemic patients to balance intake
of folic acid, cobalamin, and iron

Vitamin C (aka ascorbic acid)

- Found in citrus fruits, guava, kiwi, broccoli, brussel
sprouts
- Coenzyme:
o N/A ascorbic acid acts as a cofactor
o Most powerful reducing agent known to occur naturally in
living tissue
o Acts as cofactor in the hydroxylation of proline to
hydroxyproline (creating more OH
groups for H bonding) which gives the
triple helix of collagen its strength
o Ascorbic acid is important due to its
ability to maintain metal ions (also cofactors) in the correct
ionic state within an enzyme’s active site (e.g. Fe2+ in the
active site of prolyl hydroxylase)

38
 o Acts as cofactor in steroid synthesis, noradrenaline
production and the conversion of folic acid to THF
o Important independent role within
body as an antioxidant, donating
electrons to limit damage caused by
free radicals and oxidative species
o Also involved in immune system regulation (especially during
infection)
- Deficiency state:
o Scurvy is characterised by swollen gums, bruising,
haemorrhage, bone fracture, loose teeth, poor wound
healing, anaemia
o Supplements may benefit those not following a balanced diet,
diabetics, pregnant/lactating women, heavy drinkers, and
smokers
- Toxicity state:
o Stomach complaints

Fat Soluble Vitamins

Vitamin A (aka retinol)
- Found in cod liver oil, butter, milk, cheese
- Retinol can be synthesised from
carotenoids (found in carrots and tomatoes)
- Coenzyme:
o N/A retinol does not form a coenzyme
o Retinol is converted into retinal and retinoic
acid in the body
o Retinal is further converted to I I-cis-retinal
(using NADP+ as coenzyme) which is a
photosensitive prosthetic group essential for
normal function of retina
o The retinal molecule undergoes isomerisation
in presence of a photon of the correct wavelength of light

39
 o It is regenerated via the RPE65 enzyme so that it can be used
to detect light again
o Retinoic acid is required for growth, immune
system function, maintenance of epithelial
tissue (including skin), normal embryonic
development
- Deficiency state:
o Still widespread in developing world
o Symptoms include night blindness, dry/hyperkeratotic skin,
skin infection, corneal damage, blindness
o Supplements may be required in breast feeding women to
compensate for infant’s supply and in patients unable to
absorb or store lipids
- Toxicity state:
o Toxicity to unborn children (C/I using retinoic acid in
pregnancy)
o Hypervitaminosis A – excessive intake can lead to liver
damage, bone pain, vision changes and death

Vitamin D (aka calciferol)

- Ergocalciferol is found in fungi sources (e.g. mushrooms)
- Cholecalciferol is found in animal sources
(e.g. fatty fish, cod liver oil, egg yolk)
- Body not entirely dependent on dietary
intake as vit D3 is formed under UV light
from provitamin precursors (7-dehydrocholesterol) in the skin
- Coenzyme:
o N/A calciferol does not form a coenzyme
o Calciferol is converted into calcifediol
in the liver before being further altered
in the kidney to produce calcitriol
o Calcitriol stimulates synthesis of
specific proteins that act as Ca2+ carriers in bone and
intestine increasing absorption of dietary Ca2+ and release of

40
 Ca2+ from bone (PO4- passively accompanies the Ca2+
movements)
o Calcitriol maintains Ca2+ and PO4- at sites of new bone
formation therefore is essential for proper formation of the
skeleton
- Deficiency state:
o Leads to rickets in children characterised by distortion of the
long bones of the legs and bones of the pelvis and spine
o In adults it leads to osteomalacia (bone softening)
o Rate of synthesis of vit D in the skin depends on exposure to
UV light therefore night workers and miners traditionally
prone to deficiency
o Rate of synthesis of vit D in skin also depends of skin
pigmentation
o Melanin protects skin cells by preventing UV light penetration
into the skin, this reduces the efficiency of the conversion of
7-dehydrocholesterol
- Toxicity state:
o Hypercalcaemia – calcium deposits in organs (e.g. kidneys,
liver, heart), anorexia, insomnia, abnormal bone formation

Vitamin E (aka the tocopherols and tocotrienols)

- Found in vegetable oils
- Coenzyme:
o N/A do not form coenzymes
o Act as a fat-soluble antioxidant
o Particularly important in protecting cell
membranes from oxidative damage by
free radical species
o Also involved in smooth muscle growth and maintenance of
nerves
- Deficiency state:
o Usually due to malabsorption of fat rather than lack of dietary
intake

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 o Symptoms include nerve damage and haemolytic anaemia
- Toxicity state:
o Antagonises vit K leading to risk of bleeding

Vitamin K (aka phytomenadione (K1), menaquinone (K2))

- Leafy green vegs (e.g. spinach, cabbage,
kale) contain high levels of
phytomenadione
- Menaquinone is found in eggs, meat, and
dairy
- Intestinal flora can convert phylloquinone into menaquinone
- Coenzyme:
o N/A phytomenadione acts as a cofactor
o Acts as cofactor in reactions that add a second carboxylic acid
group onto the glutamate residues of a no. of proteins to
form a gamma-carboxyglutamate (Gla) residue
o This is important in the formation of coagulation factor II
(prothrombin), VII, IX and X in the liver
o Also involved in production of anticoagulant proteins C and S
and protein Z
o Warfarin inhibits phytomenadione’s effects on the
coagulation to mediate its clinical effect
- Deficiency state:
o Rarely due to lack of dietary intake
o Liver damage (e.g. alcoholics) and medication (e.g.
anticoagulants) can lead to deficiency
o Symptoms include anaemia, bruising and bleeding at the
mucosal membranes (e.g. gums, nose)
- Toxicity state:
o Unlike most fat-soluble vitamins, not stored in great
quantities in the liver
o Therefore, toxicity is not seen with higher doses of vit K1 or
K2 in otherwise healthy people

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Energy and Metabolism
Adenosine Triphosphate (ATP)
- Universal energy molecule
- Inside the cell in the cytoplasm
- Consists of triphosphate, deoxy-ribose
(sugar, OH) and adenine (H)
- Deoxy-ribose and adenine are one of 4
bases of DNA called “adenosine”
- Energy rich compounds: guanosine triphosphate (GTP), cytidine
triphosphate (CTP), uridine triphosphate (UTP)

- ATP contains high energy phosphate bonds

o One phosphate ester bond formed by linkage between the
alpha-phosphoryl group and the 5’-oxygen of ribose
o Phosphoanhydride bonds
formed alpha, beta and beta,
gamma linkages between
phosphoryl groups
o Usually present as complex
magnesium cations

Overview of Catabolic Processes

- Metabolism is divided into catabolism (producing energy) and
anabolism (producing molecules)
- The main route is:
o Simple sugars go through glycolysis where they are broken
down and pyruvate is produced with a small amount of ATP
o Pyruvate is converted into acetyl CoA
o Acetyl CoA enters the citric acid cycle (Krebs cycle) where CO2
and e- are produced
o E- are transported to the oxidative phosphorylation stage
where ATP is produced
- Fats are broken down into glycerine and that enters the catabolic
routes at the glycolysis stage

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 - Fatty acids are broken and enter catabolism as acetyl CoA
- Proteins are broken down into amino acids and depending on the
amino acid, they enter catabolism in different places

- Stage 1 – no useful energy generated

- Stage 2 – small amount of ATP generated – anaerobic
- Stage 3 – majority of ATP production – aerobic

Energy Supply
- For most organisms the main supply of energy is by metabolism of
glucose and other sugars
- Metabolism of fats and proteins becomes more important for
energy production when the supply of glucose and other sugars is
limited
- The initial stage in the metabolism of glucose is called glycolysis

Glycolysis
- Takes place in the cytoplasm
- Does not need oxygen

44
 - Not efficient
- Produces 2 molecules of NADH (nicotinamide adenine dinucleotide)
- Uses energy to produce more ATP
- Gives starting point for other stages

- Divided into 2 parts

1. 6 carbon units which required energy (hexose stage)
2. 3 carbon units which provides energy (triose stage)

- Consists of 10 different reactions (6 different reaction types)

o Phosphoryl transfer
o Phosphoryl shift
o Isomerisation
o Dehydration
o Aldol cleavage
o Oxidation
- Uses 10 different enzymes

- Consumption/Generation of ATP
o Glucose  glucose 6-phosphate -1
o Fructose 6-phosphate  fructose 1,6-bisphosphate -1
o 2x 1,3-bisphosphoglycerate  2x 3-phosphoglycerate +2
o 2x phosphoenol pyruvate  2x pyruvate +2
o Overall = +2

- Process
1. Glucose phosphorylated by ATP to form G6P [hexokinase]

45
 2. G6P converted to fructose-6-phosphate [glucose isomerase]
reversible reaction
3. Fructose-6-phosphate phosphorylated by ATP to form fructose
1,6-bisphosphate [phosphofructokinase]
4. Fructose 1,6-bisphosphate converted into GA3P + DHAP
[aldolase] reversible reaction
5. DHAP is isomerized to form a 2nd GA3P molecule [triose
phosphate isomerase] reversible reaction
6. The 2 GA3P molecules are oxidised using NAD+ to form 1,3-BPG
[glyceraldehyde 3-phosphate dehydrogenase] reversible
reaction, produces NADH
7. 1,3-BPG converted to 3PG [phosphoglycerate kinase] reversible
reaction, produces ATP
8. 3PG converted to 2PG [phosphoglycerate mutase] reversible
reaction
9. Water removed from 2PG to form phosphoenolpyruvate
[enolase] reversible reaction
10. Phosphoenolpyruvate converted to pyruvate [pyruvate
kinase] produces ATP

NAD+/NADH
- NADH is the reduced form of NAD+
- NAD+ is the oxidised form of NADH
- NAD+
- READ ABOUT structure/synthesis/use of NAD+/NADH

Fate of Pyruvate
- Conversion to acetyl CoA
o Used in citric acid cycle and electron transport chain
o Formation of acetyl CoA occurs in the
mitochondria
- Conversion to lactate
- Conversion to ethanol

46
 - READ ABOUT structure/synthesis/use acetyl CoA

Citric Acid Cycle

- Aka tricarboxylic acid cycle or Krebs cycle
- Occurs in mitochondria
- Requires oxygen
- Entry point is acetyl CoA
- Acetyl CoA is fully oxidised to carbon dioxide and water
- Also provides intermediates

- Does not produce any ATP itself

- Produces NADH and FADH2 which are converted into ATP during
the electron transport chain
- 3 molecules of NADH
- 1 molecule of FADH2 (flavin adenine dinucleotide)
- 1 molecule of GTP
- GTP is a high energy molecule
- READ ABOUT structure/synthesis/use FAD/FADH2

47
 - Process
1. Acetyl CoA joins with oxaloacetate to form citrate
2. Citrate converted to isocitrate (isomer of citrate)
3. Isocitrate is oxidised to α-ketoglutarate which results in CO2
release and formation of NADH [isocitrate dehydrogenase] rate-
limiting step
4. Α-ketoglutarate is oxidised to form a 4C molecules which binds
to CoA to form succinyl CoA. This produces another NADH, and
CO2 is released
5. Succinyl CoA converted to succinate and one GTP is produced
6. Succinate is converted to fumarate and one FADH2 is produced
7. Fumarate is converted to malate using water
8. Malate is converted to oxaloacetate and 3rd NADH is produced

Overall Stoichiometry
- Acetyl CoA + 3NAD+ + FAD + GDP + Pi + 2H2O
- 2CO2 + 3NADH + FADH2 + GTP + 2H+ + CoA
- While it doesn’t directly involve oxygen, regeneration of NAD+ and
FAD requires molecular oxygen (see oxidative phosphorylation)
- Citric acid cycle only works under aerobic conditions

Biosynthetic Intermediate

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Summary
- Glycolysis
o Converts glucose to 2 molecules of pyruvate
o Produces 2 ATP and 2 NADH
- Pyruvate to acetyl CoA
o Produces 2 NADH (per glucose)
- Citric acid cycle
o Converts acetyl CoA to CO2
o Produces 2 GTP, 6 NADH, 2 FADH2

Reading
- Gerald Karp – Cell Biology

Oxidative Phosphorylation
- Occurs in mitochondria
- NADH and FADH2 react with oxygen
- This releases energy which makes ATP

49
 - Each molecule of NADH produces 3 ATP
- Each molecule of FADH2 produces 2 ATP
- Each molecule of glucose when metabolised gives rise to 36 ATP
- 32 of these are formed in oxidative phosphorylation

- During OP, protons (H+) are pumped out of the mitochondrial

matrix
- This gives a proton gradient
mMaatrix
trix
(energy gradient)
- The gradient is used by ATP N AD H
H + H2 O AD P ATP
synthesise enzyme to O2
– – – –
synthesise ATP from ADP
- The return of protons to the H+ H+ H+ H+
matrix is couple to H+
phosphorylation of ADP

Electron Transport Chain

- Electrons are transferred from NADH (or FADH2) to molecular
oxygen by the ETC
- In the ETC there are 4 protein complexes
1. NADH-ubiquinone oxidoreductase (Complex I)
2. Succinate-ubiquinone oxidoreductase (Complex II)
3. Ubiquinol-cytochrome c oxidoreductase (Complex III)
4. Cytochrome c oxidase (Complex IV)
- There are also 2 electron carriers
o Ubiquinone
o Cytochrome c

1. NADH gives protons to complex I and becomes NAD+

2. Protons move from complex I+H to complex II which becomes
complex II+H
3. This carries on

50
 4. In the end, proton is carried to oxygen and then becomes H2O

- Succinate is converted to fumarate, taking H

- Fumarate goes back to the Krebs cycle

- So 3 protein complexes (NADH-Q reductase, cytochrome reductase,

cytochrome oxidase) have pumped protons out of the matrix of the
mitochondrion
- This generates a proton motive force
- Protons flow back into the matrix through ATP synthase (complex
V) which couples ADP and phosphate (ATP generated)

ATP Synthase
- Contains a transmembrane region and a large head
group on the matrix side of the membrane
- The head group contains the ATP synthesising
domain
- The transmembrane region is the proton channel
through which the protons flow

Mitochondria Structure
- Outer membrane
o Binds the mitochondrion
o Permeable to most small
molecules
- Inner membrane
o Highly folded
o Virtually impermeable to all
ions and polar molecules
o Contains specific transport
proteins for some molecules
such as ADP
o Site of oxidative phosphorylation
- Matrix

51
 o Bounded by inner membrane
o Site of citric acid cycle and fatty acid oxidation

Energy Sources
- Carbohydrates
o Sucrose  glucose + fructose
o Maltose  glucose
o Starch  glucose
- Lipids
o Triglycerides  glycerine + fatty acids

- Proteins

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