BIOCHEMISTRY – LECTURE: 1ST YEAR 2ND SEMESTER MIDTERM

COVERAGE 2. Type of reaction catalyzed by an enzyme is often used as a

prefix
1. Enzymes and Vitamins
 E.g., Oxidase - catalyzes an oxidation reaction,
2. Nucleic Acids
 E.g., Hydrolase - catalyzes a hydrolysis reaction
3. Digestion
3. Identity of substrate is often used in addition to the type of
4. Bioenergy Production reaction
 E.g. Glucose oxidase, pyruvate carboxylase, and succinate
ENZYMES AND VITAMINS dehydrogenase
- Enzymes are catalysts and are not consumed in the reactions
- Enzymes are proteins that act as a catalyst for biochemical Six Major Classes
reactions - Enzymes are grouped into six major classes based on the types
- The human body has 1000s of enzymes of reactions they catalyze
- Enzymes are the most effective catalysts known Class Reaction Catalyzed
- Most enzymes are globular proteins 1. Oxidoreductases Oxidation-reductions
- A few enzymes are now known to be ribonucleic acids (RNA)
2. Transferases Functional group transfer reactions
- Enzymes undergo all the reactions of proteins including
3. Hydrolases Hydrolysis reactions
denaturation
4. Lyases Reactions involving addition or removal
- Enzyme activity is dramatically affected by:
of groups form double bonds
 Alterations in pH
5. Isomerase Isomerisation reactions
 Temperature
6. Ligases Reactions involving bond formation
 Other protein denaturants
coupled with ATP hydrolysis
Simple and Conjugated Enzymes
Oxidoreductase
 Simple enzyme: composed only of protein (amino acid chains) - catalyzes an oxidation–reduction reaction:
 Conjugated enzyme: Has a nonprotein part in addition to a  Oxidation and reduction reactions are always linked to one
protein part. another
o Apoenzyme: Protein part of a conjugated enzyme.
 An oxidoreductase requires a coenzyme that is either
o A cofactor: Nonprotein part of a conjugated enzyme.
oxidized or reduced as the substrate in the reaction.
o A holoenzyme is the biochemically active conjugated
 E.g., Lactate dehydrogenase is an oxidoreductase and the
enzyme
reaction catalyzed is shown below
o Apoenzyme + cofactor = holoenzyme (conjugated enzyme)
Transferase
- an enzyme that catalyzes the transfer of a functional group from
Cofactors
one molecule to another
- important for the chemically reactive enzymes
- Two major subtypes:
- small organic molecules or Inorganic ions
- Organic molecule cofactors: also called as co-enzymes or co-  Transaminases - catalyze transfer of an amino group to a
substrates substrate
- Co-enzymes/co-substrates are derived from dietary vitamins  Kinases - catalyze transfer of a phosphate group from
- Inorganic ion cofactors adenosine triphosphate (ATP) to a substrate
- Typical metal ion cofactors - Zn2+, Mg2+, Mn2+, and Fe2+ Hydrolase
- Nonmetallic ion cofactor - Cl- - A hydrolase is an enzyme that catalyzes a hydrolysis reaction
- Inorganic ion cofactors derived from dietary minerals - The reaction involves addition of a water molecule to a bond to
cause bond breakage
Nomenclature and Classification of Enzymes - Hydrolysis reactions are central to the process of digestion:
- Nomenclature: Most commonly named with reference to their  Carbohydrase’s hydrolyze glycosidic bonds in oligo- and
function polysaccharides (see reaction below)
 Type of reaction catalyzed  Proteases effect the breaking of peptide linkages in proteins,
 Identity of the substrate  Lipases effect the breaking of ester linkages in
- A substrate is the reactant in an enzyme-catalyzed reaction: triacylglycerols
 The substrate is the substance upon which the enzyme Lyase
“acts.” - an enzyme that catalyzes the addition of a group to a double
 E.g., In the fermentation process sugar to be converted to bond or the removal of a group to form a double bond in a
CO2, therefore in this reaction sugar is the substrate manner that does not involve hydrolysis or oxidation
 Dehydratase: effects the removal of the components of
Three Important Aspects of the Naming Process water from a double bond
1. Suffix -ase identifies it as an enzyme  Hydratase: effects the addition of the components of water
 E.g., urease, sucrase, and lipase are all enzyme to a double bond
designations Isomerase and Ligase
 Exception: The suffix -in is still found in the names of some - An isomerase is an enzyme that catalyzes the isomerization
digestive enzymes, E.g., trypsin, chymotrypsin, and pepsin (rearrangement of atoms) reactions.

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 BIOCHEMISTRY – LECTURE: 1ST YEAR 2ND SEMESTER MIDTERM

Introns and Exons Characteristics of the Genetic Code

- In prokaryotes, each gene is a continuous segment along a DNA - The genetic code applies almost universally: with minor
molecule. exceptions, the same amino acid is represented by the same
- Transcription of the gene produces mRNA that is translated into codon(s) in all species.
a protein almost immediately, because there is no nuclear - Most amino acids are represented by more than one codon (a
membrane separating the DNA from the cytoplasm. feature known as degeneracy).
- In eukaryotes, the gene segments of DNA that code for proteins  Only methionine and tryptophan are represented by a single
(exons) are interrupted by segments that do not carry an amino codon.
acidcode (introns).  Leucine, serine, and arginine are represented by six codons.
- Both exon and intron segments are transcribed, producing  No codon codes for more than one amino acid.
heterogenous nuclear RNA (hnRNA). - Only 61 of the 64 possible triplets represent amino acids. The
- A series of enzymes cut out the intron segments and splice the other three are used as signals for chain termination (a “stop”
exon segments together to produce mRNA. signal).
- The AUG codon (which also codes for methionine) functions as
a “start” signal, but only when it occurs as the first codon in a
sequence.

Genetic Code
- Once the 3D structure of DNA was known, it was clear that the
sequence of the bases along the backbone in some way directed Translation and Protein Synthesis
the order in which amino acids were stacked to make proteins. - Several ribosomes can move along a single strand of mRNA,
- In 1961, Marshall Nirenberg and his coworkers began to producing several identical proteins simultaneously. These
unravel the connection between the base sequence in DNA and complexes are called polyribosomes or polysomes.
the amino acid sequence in proteins. - The growing polypeptide chain emerging from the end of the
- The genetic code uses a sequence of three bases (a triplet code) ribosome spontaneously folds into the characteristic 3D shape
to specify each amino acid. (A triplet code gives 43=64 possible of that protein.
combinations, which is more than enough to specify the 20 Step 1: Initiation of the polypeptide chain.
amino acids.) - mRNA and a small ribosomal subunit join; the initiating codon
- Each base triplet sequence that represents a code word on Mrna (AUG) is aligned with P (peptidyl) site of the subunit.
molecules is called a codon. - tRNA brings in methionine (eukaryotes) or N-formyl
methionine (prokaryotes).
- The resulting complex binds to the large ribosomal subunit to
form a unit called the initiation complex.
Step 2: Elongation of the chain.
- The next incoming tRNA with an anticodon that is
complementary to the mRNA codon bonds at the A (aminoacyl)
site on the mRNA.
- A peptide bond is formed between the amino acid segments,
(catalyzed by peptidyl transferase), which releases the amino
acid chain from the P site.
- The “empty” tRNA released, and the whole ribosome moves
one codon along the mRNA towards the 3’ end (translocation).
- Another tRNA attaches to the A site, and the elongation process
is repeated.
Step 3: Termination of polypeptide synthesis.
- Elongation continues until the ribosome complex reaches a stop
codon (UAA, UAG, or UGA).
- A termination factor protein binds to the stop codon, and
separates the protein from the final tRNA.
- The ribosome can then synthesize another protein molecule.

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 BIOCHEMISTRY – LECTURE: 1ST YEAR 2ND SEMESTER MIDTERM

Reactions of the Citric Acid Cycle - Several intermediate reactions are involved in this electron
 Step 1: Formation of Citrate transfer
 Step 2: Formation of Isocitrate Complex II: Succinate-coenzyme Q Reductase
 Step 3: Oxidation of Isocitrate and Formation of CO2: involves - Smaller than complex I
oxidation–reduction as well as decarboxylation - Contains only four subunits including two iron-sulfur protein
 Step 4: Oxidation of Alpha-Ketoglutarate and Formation of clusters (FeSP)
CO2 - Succinate is converted to fumarate by this complex
 Step 5: Thioester bond cleavage in Succinyl CoA and - In the process it generates FADH2
Phosphorylation of GDP - CoQ is the final recipient of the electrons from FADH2
 Step 6: Oxidation of Succinate Complex III: Coenzyme Q – Cytochrom c Reductase
- Complex III contains 11 different subunits
 Step 7: Hydration of Fumarate
- Several iron-sulfur proteins and cytochomes are electron
 Step 8: Oxidation of L-Malate to Regenerate Oxaloacetate
carriers in this comple
Regulation of the Citric Acid Cycle
- Cytochrome is a heme iron protein in which reversible
- The rate at which the citric acid cycle operates is controlled by
oxidation of an iron atom occurs
ATP and NADH levels
- Various cytochromes, e.g., cyt a, cyt b, cyt c, differ from each
- When ATP supply is high, ATP inhibits citrate synthase (Step
other by:
1 of Citric acid cycle)
- When ATP levels are low ADP, ADP activates citrate synthase  Their protein constituents
- Similarly, ADP and NADH control isocitrate dehydrogenase:  The manner in which the heme is bonded to the protein
 NADH acts as an inhibitor  Attachments to the heme ring
Complex IV: Cytochrome c Oxidase
 ADP as an activator.
- Contains 13 subunits including two cytochromes
Electron Transport Chain - The electrons flow from cyt c to cyt a to cyt a3
- In the final stage of electron transfer, the electrons from cyt a3,
- The electron transport chain (ETC) facilitates the passage of
and hydrogen ion (H+) combine with oxygen (O2) to form
electrons trapped in FADH2 and NADH during citric cycle
- ETC is a series of biochemical reactions in which intermediate water
carriers (protein and non-protein) aid the transfer of electrons
and hydrogen ions from NADH and FADH2 - It is estimated that 95 % of the oxygen used by cells serves as
- The ultimately receiver of electrons is molecular oxygen the final electron acceptor for the ETC
- The electron transport (respiratory chain) gets its name from the
fact electrons are transported to oxygen absorbed via respiration Oxidative phosphorylation
- The overall ETC reaction: - process by which ATP is synthesized from ADP and Pi using
the energy released in the electron transport chain. - coupled
- Energy is used to synthesize ATP in oxidative phosphorylation reactions
- Note that 2 hydrogen ions, 2 electrons, and one half-oxygen - Coupled Reactions – are pairs of biochemical reactions that
molecule react to form the product water occur concurrently in which energy released by one reaction is
- This relatively straight forward reaction actually requires eight used in the other reaction
or more steps  Example: oxidative phophorylation and the oxidation
- The reaction releases energy (exothermic reaction) reactions of the electron transport chain are coupled
- The energy released is coupled with the formation of three ATP systems
molecules per every molecule of NADH processed through - The coupling of ATP synthesis with the reactions of the ETC is
ETC related to the movement of protons (H+ ions) across the inner
- The enzymes and electron carriers needed for the ETC are mitochondrial membrane
located along inner mitochodrial membrane - Complexes I, III and IV of ETC chain have a second function
- They are organized into four distinct protein complexes and two in which they serve as “proton pumps” transferring protons
mobile carriers from the matrix side of the inner mitochondrial membrane to
- The four protein complexes tightly bound to membrane: the intermembrane space
 Complex 1: NADH-coenzyme Q reductase - For every two electrons passed through ETC, four protons cross
the inner mitochondrial membrane through complex I, four
 Complex II: Succinate-coenzyme Q reductase
through complex III and two more though complex IV
 Complex III: Coenzyme Q - cytochrome C reductase
- This proton flow causes a buildup of H+ in the intermembrane
 Complex IV: Cytochrome C oxidase space
- Two mobile electron carriers are: - The gradient build-up would push the H+ ions through
 Coenzyme Q and cytochrome c. membrane-bound ATP synthase:
Complex 1: NADH-Coenzyme Q Reductase  This high concentration of protons passing through ATP
- NADH from citric acid cycle is the source of electrons for this synthase becomes the basis for the ATP synthesis
complex
- It contains >40 subunits including flavin mononucleotide
(FMN) and several iron-sulfur protein clusters (FeSP)
- Net result: Facilitates transfer of electrons from NADH to
coenzyme Q
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