PBI1092 Academic English 2

Semester 1, 2019/2020

Assessment 2b: Explanation Report

Title: Citric Acid Cycle (Krebs Cycle)

Group: G23

Instructor: Mr. Mohamad Fairuz bin Ali

Group Members Matric Number

1. Anis Jasmin Binti Azimin 72116
2. Neilveen Gima Anak Matin 70714
3. Anis Sofia Binti Azmi 72118
4. Siti Humairah Binti Shabri 65210
 The Krebs cycle, also known as the citric acid cycle or the tricarboxylic acid
cycle, is one of the most important reaction sequences in biochemistry. Not only is
this series of reactions responsible for most of the energy needs in complex organisms,
the molecules that are produced in these reactions can be used as building blocks for a
large number of important processes, including the synthesis of fatty acids, steroids,
cholesterol, amino acids for building proteins (Cleveland & Morris, 2014). There
were eight steps of Krebs Cycle proposed by Krebs and Johnson (1937) and was cited
and edited by Plowman and Smith (2006) are formation of citric acid, formation of
isocitrate, formation of alpha-ketoglutarate, formation of succinyl – CoA, generation
of succinate, formation of fumarate, formation of malate and regeneration of
oxaloacetate (Refer to Appendix 1).

The first step of Krebs cycle is formation of citric acid, acetyl CoA reacts with
oxaloacetate and water to form citrate, HS–CoA and a proton (Horton, Moran, Perry
& Scrimgeour, 2012). The Citrate synthase links to the oxaloacetate substrate which
can then bind to the acetyl-CoA’s acetyl group, which then releases the co-enzyme A.
This produces the very familiar and common citric acid (Horton et al., 2012). Citrate
synthase undergoes a large conformational change on binding oxaloacetate. Horton et
al. (2012) found that this binding site lies at the base of a deep cleft between the small
domain of one subunit and the large domain of the other subunit. When oxaloacetate
is bound, the small domain rotates and this closure creates the binding site for
acetyl-CoA which is a site formed by amino acid side chains from both small and
large domains. When the reaction is complete, coenzyme A is released. The enzyme
then reverts to the open conformation when citrate is released (Horton et al., 2012).

The second step of Krebs cycle is the formation if isocitrate. Aconitase catalyses
a near-equilibrium conversion of citrate to isocitrate (Horton et al., 2012). Citrate is a
tertiary alcohol and thus cannot be oxidized directly to keto acid. The formation of
keto acid intermediate is required for the oxidative carboxylation reaction that occurs
in step 3 of the citric acid cycle. As discussed by Horton et al. (2012) the step
catalysed by aconitase creates a secondary alcohol in preparation of step 3. So, the
aconitase links to the citrate to move one of its oxygen atoms to create a more
unstable citrate isomer. It does this by extracting a water molecule producing
cis-aconitate and then reattaching the water molecule to produce isocitrate (Horton et
al., 2012).

The third step of Krebs cycle is formation of alpha-ketoglutarate where isocitrate

dehydrogenase catalyses the oxidative decarboxylation of isocitrate to form
a-ketoglutarate (Horton et al., 2012). This reaction is the first of four
oxidation-reduction reactions in citric acid cycle. Horton et al. (2012) also stated that
when the citrate rearranged, the process begins in earnest; the isocitrate
dehydrogenase links to the isocitrate, which then transfers an electron to the NAD+,
Nicotinamide adenine dinucleotide, producing its energized form NADH. With the
electron removed, the enzyme then detaches a carbon atom to form a molecule of
carbon dioxide. This transforms the substrate from a 6-carbon molecule to a 5-carbon
molecule (Horton et al., 2012).

The fourth step of Krebs cycle is formation of succinyl – CoA, subsequent

conversion of α-ketoglutarate to succinate is mediated via the thioester, succinyl-CoA.
Succinyl-CoA is first formed by decarboxylation and reduction of α-ketoglutarate,
with the amalgamation of CoA, catalysed by the α-ketoglutarate dehydrogenase
complex (Clarke, 2013). The next two steps are catalyzed by isocitrate
dehydrogenase. Dehydrogenation of isocitrate forms oxalosuccinate, which
decarboxylates to alpha-ketoglutarate. Alpha-ketoglutarate is further oxidatively
decarboxylated by alpha-ketoglutarate dehydrogenase—a multienzyme
complex. Succinyl-CoA is formed in this unidirectional reaction (Kumari, 2018).

The fifth step of Krebs cycle is generation of succinate The succinyl-CoA is

converted to succinate by the enzyme succinyl-CoA synthase and substrate-level
phosphorylation takes place with the slight twist that GDP is the phophate acceptor
(Dunford & Doyle, 2007). Dunford and Doyle (2007) mention that the breakdown of
succinyl coenzyme A is coupled to the phosphorylation of GDP to form guanosine
triphosphate, GTP. Scheffler (n.d.) stated that the inorganic phosphate displaces the
CoA to form succinyl-phosphate can be proved from isotope tracer experiments. This
is the only step in the Krebs cycle that produces and stores energy directly and as in
glycolysis, this ATP is produced by subsrate-level phosphorylation (Plowman &
Smith, 2006).
 The sixth step of Krebs cycle is formation of fumarate. Succinate is converted
into fumarate by the membrane-linked enzyme succinate dehydrogenase (SDH). As
discussed by Scheffler (n.d.) succinate is oxidised when two of its hydrogen are
transferred to FAD forming FADH2 where a large flavoprotein forms the active site,
and a covalently linked flavin becomes the hydrogen acceptor. The flavin is linked to
a histidine side chain in a highly conserved portion of the protein (Scheffler, n.d.). It
is immediately reoxidized with the release of protons and the transfer of two electrons
to the iron-sulfur centers of the iron protein subunit of SDH. Succinate dehydrogenase
is the only Krebs cycle enzyme directly linked to the electron transfer chain (Scheffler,
n.d.).

The seventh step of Krebs cycle is the formation of malate. This process includes
the catalyzation process of fumarate by the enzyme named fumarase. This reaction
involves the addition of water molecules. Fumarate is a prochiral molecule
(Damodaran, Kannan & Sreekumari, 2017). Horton et al. (2012) found that when
fumarate is positioned in the active site of fumarase, the double bond of the substrate
can be attacked from only one direction. The product of the reaction is exclusively the
L stereoisomer of the hydroxy acid malate. There are two unrelated fumarases that
can catalyze the same reaction. The class I enzyme is found in most bacteria. The
class II enzyme is present in some bacteria and all eukaryotes. Some bacteria, such as
E. coli, have both forms of the enzyme. One form is active in the normal citric acid
cycle pathway and the other usually specializes in the reverse reaction to convert
malate to fumarate (Horton et al., 2012).

The last step in the Krebs Cycle is the degeneration of oxaloacetate process
which involve the oxidization process of malate by malate dehydrogenase (Horton et
al., 2012). The coenzyme is NAD+. This reaction is catalyzed by NAD+ dependent
malate dehydrogenase. The near-equilibrium interconversion of the a-hydroxy acid
L-malate and the keto acid oxaloacetate is analogous to the reversible reaction
catalyzed by lactate dehydrogenase. Horton et al. (2012) further claimed that since
this is a near-equilibrium reaction it means that under the conditions found inside the
cell, the concentration of malate is very much higher than that of oxaloacetate and the
NADH is generated. The oxaloacetate can further condense with another acetyl -CoA
molecule and the cycle continues (Horton et al., 2012).
References

Clarke, K. G. (2013). Bioprocess Engineering: An Introductory Engineering and Life

Science Approach. Cambridge, United Kingdom: Elsevier Science and
Technology.

Cleveland, C. J. & Morris C. G. (2014). Dictionary of Energy. London, United

Kingdom: Elsevier Health Science.

Damodaran, V., Kannan, V., & Sreekumari S. (2017). Citric acid cycle. Textbook of
Biochemistry for Medical Students (pp. 303-311).

Dunford, M., & Doyle, A. (2007). Energy systems and exercise. Nutrition for Sport
and Exercise (pp. 71-72). United State of America: Peter Adams.

Horton, R., Moran, L. A., Perry, M. D., & Scrimgeour, K. G. (2012). Principles of
biochemistry. The Citric Acid Cycle (5th ed.) ( pp. 385-416).

Kumari, A. (2017). Sweet Biochemistry: Remembering Structures, Cycles, and

Pathways by Mnemonics. San Diego, United States: Elsevier Science Publishing
Co Inc.

Plowman, S. A., & Smith, D. L. (2006). Exercise Physiology for Health, Fitness and
Performance. Philadelphia, Pennsylvania: Lippincott Williams & Wilkins.

Scheffler, I. E. (n.d.). Metabolic pathways inside mithochondria. Mitochondria (pp.

250). San Diego, California: John Wiley & Sons.
Appendix 1

Figure 1. Citric Acid Cycle (Krebs Cycle)