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Файл:Carbohydrates. Bioenergetics. Study manual
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enzyme. Write a balanced equation for the reaction catalyzed by this enzyme. Explain how ATP can be a substrate and inhibitor of phosphofructokinase? How is the activity of this enzyme through ATP? How glycolysis is regulated depending on the level of the ATP?
Task 4. Glycogen phosphorylasefrom skeletal muscle is characterized by a
much higher magnitude of V
than the same enzyme from liver tissue. What
max
physiological function does glycogen phosphorylasein skeletal muscle and liver tissue? Why the magnitude of V
for the muscle enzyme need to be greater than for
max
the enzyme from the liver?
Task 5. Is it possible a real synthesis of glucose from pyruvate under conditions
when the citric acid cycle and oxidative phosphorylation is completely inhibited? Explain your answer.
Task 6. Liver extract capable of catalyzing all the normal metabolic reactions
are incubated in separate experiments with the following precursors labeled14C:
а) НО--14СОО-– bicarbonate
б) СН3--CО--14СОО-– pyruvate.
Trace the path of each of these two precursors in gluconeogenesis. In what position is found mark all the intermediate products and the final product, i.e. glucose?
Task 7. How does increasing the concentration of ATP and AMP on the catalytic
activity of fructose diphosphatasesand phosphofructokinase? As the impact of these
effects on the value flows of metabolites of gluconeogenesis and glycolysis? Why?
Task 8. To determine whether or not a compound to serve as a precursor of glucose, typically in the following manner: the animal is left to starve until he depleted
the glycogen stores, and then give him the test compound. Those connections, which
influences the amount of glycogen in the liver is increased, are called glucogenic,
because, first, they are transformed into glucose-6-phosphate. Below are the names
of some compounds. On the basis of known enzymatic reactions, indicate which
ones areglucogenic: a) succinate, b) glycerol, C). acetyl-CoA, g) pyruvate d) butyrate. Give a scheme of synthesis of glucose from the chosen compounds.
Task 9.
The level of lactate in the blood during exercise.
The figure shows the concentration of lactate in the blood to race in the 400 m
and beyond. What caused the rapid rise in lactate concentration? What is the reason
for the decrease in the level of lactate after the race? Confirm your explanation with
a diagram. Why the decline is slower than the rise? Why at rest the concentration of
lactate in the blood is not equal to zero?
Task 10. Calculate how many ATP molecules formed during the conversion of
glucose to 1,3-diphosphoglycerate in aerobic conditions.
Task 11. Calculate how many ATP molecules required for the synthesis in the
liver of one molecule of glucose from lactate. How to change the synthesis of glucose in the reduce the ratio of ATP/ADP? Explain why.
Task 12. In some diseases (e.g., malignant neoplasms of the pancreas), increased synthesis of insulin. Patients you experience increased hunger, increased fatigue, weakness. In the future, joins brain injury. Why develop the described symptoms? What is the mechanism of the observed brain damage?
Problem 13. If the fraction of membranes precipitated by centrifugation of the
homogenate of the liver was separated and treated with adrenaline, observed the
formation of a new substance. This substance was isolated and purified. Unlike
adrenaline when added to the supernatant fluid of the homogenate it activated glycogen phosphorylase. Explain the obtained data.
Problem 14. The addition of epinephrine to a homogenate or drug damaged
cells to a healthy liver lead to increase in glycogen phosphorylase. However, if the
homogenate was pre-centrifuged at high speed and then the clear supernatant fluid
was added adrenaline or glucagon, the increase phosphorylase activity was observed. Explain the results obtained.
Task 15. Under stress the release of adrenaline stimulates the breakdown of
glycogen in the liver, heart and skeletal muscle. A product of the breakdown of glycogen in the liver is glucose in skeletal muscle glycogen is broken down in the
course of glycogenolysis. Why final products of the breakdown of glycogen in these
tissues are different?
Problem 16. How many molecules of ATP is formed in aerobic conditions the
oxidation of one glucose molecule to pyruvic acid? How to change energy effect, if
the oxidation of glucose will occur in anaerobic conditions? Reasons for the answer.
Problem 17. What are the isoforms of lactate dehydrogenase (LDH) appear in
the blood of the patient:
myocardial infarction
acute hepatitis
Response to justify.
Problem 18. The patient was injected with solution of epinephrine. How to
change the level of glucose in the blood. Explain the mechanism of these changes.
Task 19. The patient discovered hyperglycemia, glucosuria, acetonemia, acetonuria. Explain the reasons for such changes, observed in the patient. What hormone is essential to the patient?
Task 20. Myocardial ischemia breaks down the process of oxidative phosphorylation, this leads to a reduction of ATP synthesis. The change of the activity of glycolysis. Response to justify.
CHAPTER 2
Bioenergy
2.1. Energy exchange
Bioenergy is a section of dynamic biochemistry that studies the regularities of
formation, accumulation and consumption of energy in biological systems. Energy exchange is a set of reactions of organic matter cleavage accompanied by energy release.
The metabolism of substances and energy, or metabolism, is a set of chemical
and physical transformations of substances and energy occurring in a living organism and providing its vital functions.
Metabolism is a set of all chemical transformations that occur in a cell or an organism and are carried out through a series of sequential enzyme-catalyzed reactions
called metabolic pathways.
Catabolism is the process of splitting organic molecules into final products. The
final products of organic matter transformations in animals and humans are CO2,
H2O and urea. Metabolites formed both in digestion and decomposition of structural
and functional components of cells are included in the processes of catabolism.
Catabolic reactions are accompanied by energy release (exergonic reactions). Catabolism of organic matter in tissues with oxygen consumption and carbon dioxide
release is called tissue respiration.
Tissue respiration stages can be represented as a scheme:
1. The stage is preparatory. At this stage, the polymers are split into monomers,
and the resulting energy is dissipated in the form of heat and is not stored. The exchange paths are called specific and reactions in the digestive tract take place.
2. The stage is oxygen-free. It occurs in the cytoplasm of the cell under anaerobic conditions (e.g. glycolysis). Intermediate products such as lactic acid, ethyl alcohol, acetic acid, acetone, etc. are formed.
3. Stage – oxygen decomposition (aerobic respiration). It takes place in mitochondria. Substances formed in the cell at the previous stages, with the participation
of oxygen decompose into the final products of CO2 and H2O. In the process of oxygen respiration, a large amount of energy is released (about 9 about 70%), which is
accumulated in the ATP molecules.
Anabolism combines biosynthetic processes in which simple building blocks
are combined into complex macromolecules necessary for the body. Anabolic reactions use energy released during catabolism (endergonic reactions). The only source
of energy for the human body is the oxidation of organic matter coming from food.
When foodstuffs are broken down to finite elements – carbon dioxide and water –
energy is released, part of which is transferred to mechanical work performed by
muscles, the other part is used to synthesize more complex compounds or is accumulated in special macroergic compounds.
2.2. Macroergic compounds
Macroergic compounds are substances whose cleavage is accompanied by the
release of large amounts of energy. Macroergic compounds include mainly adenosine triphosphoric acid (ATP) and substances capable of forming ATP in enzymatic
transport reactions of mainly phosphate groups, as well as nucleoside tri- (or di)phosphoric acids, pyrophosphoric and polyphosphoric acids, creatine phosphoric
acid, phosphopyrovinogradic acid, diphosphoglyceric acid, acetyl and succinylcoenzymes A, aminoacyl derivatives of adenylic and ribonucleic acids and others. The
energy of macroergic bonds is used to perform any work: activation of compounds
(e.g., glucose, so that the chain of its oxidative transformations can begin), synthesis
of biopolymers (nucleic acids, proteins, polysaccharides), selective absorption of
substances from the surrounding cell environment and the release of unnecessary
products from the cell, muscle contraction and restoration of the body's active state,
etc. The supply of these compounds allows the body to react quickly to changes in
external conditions and perform physical work. When a normal bond breaks down,
about 12.5 kJ/mol energy is released. The breakdown of macroergic bonds produces
energy of 25–50 kJ/mol and more. Such a connection is indicated by the symbol "~"
(squiggle, tilda). There are two macroergic bonds in ATP:
Adenosine triphosphoric acid -ATP is one of the two most important energy
sources in any cell. For example, the Na–K pump pumps 3 sodium ions out of the
cell and pumps 2 potassium ions into the cell. This current of ions is needed to main-
tain a positive charge on the outer surface of the membrane, and only with the help
of adenosine triphosphate can the channel function. The same applies to proton and
calcium channels. ATP is a precursor to the secondary cAMP (cyclic adenosine monophosphate) messenger. cAMP not only transmits the signal received by the cell
membrane receptors, but is also an allosteric effector. Allosteric effectors are substances that accelerate or slow down enzymatic reactions. For example, cyclic adenosine triphosphate inhibits enzyme synthesis, which catalyzes lactose degradation.
The adenosine triphosphate molecule itself can also be an allosteric effector.
And in such processes, ADP acts as an antagonist of ATP: if triphosphate accelerates the reaction, the diphosphate slows down, and vice versa.
Creatine is a substance of skeletal muscles, myocardium, nervous tissue. In the
form of creatine phosphate, creatine is a "depo" of macroergic bonds, used for rapid
ATP resynthesis during cellular activity.
The use of creatine phosphate for ATP resynthesis follows the scheme:
The role of creatine in muscle tissue is particularly significant. Creatine phosphate provides ATP resynthesis in the first seconds of operation, when neither anaerobic glycolysis nor aerobic oxidation of glucose and fatty acids has yet been activated, and the blood supply to the muscle has not been increased. Creatine phosphate maintains cell viability in nerve cells without oxygen. The breakdown of the
macroergic bond of creatine phosphate produces 42 kJ/mol of energy.
The macroergic bond of phosphoenolpyruvate accumulates 54 kJ/mol of energy, and the thioetheric bond (acetylcoenzymeA) accumulates 34 kJ/mol.
2.3. Biological oxidation
Biological oxidation (cellular or tissue respiration) is a set of redox reactions
occurring in cells of the body, as a result of which complex organic substances are
oxidized with oxygen with the participation of specific enzymes. Thanks to the energy of biological oxidation is provided a constant temperature of the body, sup-
ported by the synthesis of new substances, performed mechanical work, osmotic
phenomena. As a result of biological oxidation there is oxidation and removal of
toxins from the cell, neutralization of foreign compounds (xenobiotics), regulation
of metabolism.
In contrast to the known oxidation processes, biological oxidation occurs in
mild conditions, takes place in the aquatic environment, is a multistage process and
is catalyzed by enzymes whose activity is regulated. The reactions take place in
stages, and the resulting energy is stored in the form of ATP. The following types of
biological oxidation are distinguished:
1. free, at which all energy is dissipated in the form of heat rather than trans-
formed into energy of macroergic bonds;
2. Oxidation associated with ADP phosphorylation – substrate and oxidative
phosphorylation.
2.4. Tricarboxylic acid cycle (Krebs cycle)
Krebs's cycle is also called a citric acid cycle, or cellular respiration. The citric
acid cycle in living cells was discovered and studied by German biochemist Hans
Krebs, and for this work he (together with F. Lipman) was awarded the Nobel Prize
(1953). Many scientists took part in deciphering the individual reactions of this
process: A. Saint-Diegyère, A. Leninger, S.E. Severin and others.
Krebs's cycle is the final pathway of oxidation of acetyl groups, into which
most of the organic molecules that play the role of "cellular fuel" – carbohydrates,
fatty acids and amino acids – are transformed in the process of catabolism.
In one cycle, consisting of eight enzymatic reactions, a single molecule is
completely oxidized. At the Krebs cycle level, the decomposition pathways of carbohydrates, lipids and proteins are combined. Krebs cycle metabolites are used for
the synthesis of other substances (oxalic acetic acid → glucose, aspartic acid). This
cycle occurs in the mitochondria matrix.
The Krebs cycle is the main system supplying hydrogen to the mitochondrial
respiratory chain.
All pathways of catabolism are reduced to the formation of a tri-carbon compound – pyruvic acid, which is then decarboxylated by oxidative decarboxylation in
the presence of coenzyme -thiamine pyrophosphate and undergoes decarboxylation
to form acetyl-CoA. Acetyl-CoF "burns" up to two CO2 molecules in the Krebs
cycle.
1. The first reaction of the Krebs cycle is the formation of citrate – citric acid
2. In the second reaction, the formation of an isocitrate acid occurs through the
dehydration stage and the formation of cis-aconitic acid.
Note that the addition of a water molecule to cis-aconitate
is against the Markovnikov rule
3. In the third reaction, which seems to limit the speed of the Krebs cycle, the iso-
lated acid is dehydrated in the presence of NAD+-dependent isocitrate dehydrogenase:
4. In the fourth reaction, oxidative decarboxylation of α-ketoglutarate to succinyl-CoA occurs. The mechanism of this reaction is similar to the reaction of oxidative decarboxylation of pyruvate to acetyl-CoA.
5. The fifth reaction is catalyzed by the enzyme succinyl CoA synthetase. In the
course of this reaction, succinyl CoA with GDP and inorganic phosphate is converted into succinate (succinate). At the same time, the formation of high-ergic
phosphate bond GTP due to high-ergic thioether bond succinyl-CoA takes place:
6. In the sixth reaction succinate is dehydrated into fumarate. Succinate oxidation is catalyzed by succinate dehydrogenase, in the molecule of which coenzyme
FAD is covalently bound to the protein:
7. In the seventh reaction, the resulting fumarate is hydrated under the influence
of the fumarate hydrate enzyme. The product of this reaction is malate (malate). It
should be noted that fumarate hydratase has stereo specificity - in the course of this
reaction L-apple acid (malate) is formed:
8. In the eighth reaction of the tricarboxylic acid cycle, L-malate oxidation into
oxaloacetate occurs under the influence of the mitochondrial NAD-dependent
malate dehydrogenase:
The energy released as a result of oxidation of acetyl-CoA is largely concentrated in macroergic phosphate ATP bonds. Of the four pairs of hydrogen atoms,
three pairs are transferred through NAD+ to the electron transport system; in this
case, per pair, three ATP molecules are formed in the biological oxidation system
(in the process of conjugate oxidative phosphorylation), and therefore, nine ATP
molecules in total. One pair of atoms enters the system of electron transport through
FAD, – the result is the formation of 2 molecules of ATP. In the course of Krebs
cycle reactions, 1 GTP molecule is also synthesized, which is equivalent to 1 ATP
molecule. Thus, when acetyl-CoA is oxidized, 12 ATP molecules are formed in the
Krebs cycle.
Schematic representation of the Krebs cycle:
There are two key enzymes in the TCA:
1) citrate synthase (1st reaction);
2) isocitrate dehydrogenase (3rd reaction).
Both enzymes are allosterically inhibited by an excess of ATP and NADH, H+.
Isocytrate dehydrogenase is strongly activated by ADP. If there is no ADP, this enzyme is inactive. Isocytrate dehydrogenase is inhibited by ATP much more than citrate synthase, so under conditions of energy rest the concentration of citrate increases, and it is released into the cytoplasm by the concentration gradient by means
of light diffusion. In cytoplasm, citrates turn into acetyl-CoA, which is involved in
the synthesis of fatty acids. Intermediate products of Krebs cycle metabolism are
used to synthesize other substances. Amino acids are synthesized from αketoglutarate and oxaloacetate, carbohydrates are synthesized from oxalo-acetate,
and hemoglobin is synthesized from succinyl-CoA. The recovered coenzymes
NADH, H+and FADH2 in the respiratory chain are oxidized to form water, ATP and
hydrogen peroxide, a by-product.
2.5. Respiratory Chain
A set of sequential redox reactions is called an electron transfer chain or respiratory chain.
Enzymes and co-ferments in the respiratory chain.
1. NAD - dependent dehydrogenases.
As a coenzyme, they contain NAD+ and NADH. Pyridine nicotinamide ring is
able to attach electrons and hydrogen protons.
NAD-dependent dehydrogenase is located on the matrix surface of the internal
membrane of mitochondria gives a couple of hydrogen electrons to FMN-dependent
dehydrogenase. At the same time, the proton pair passes from the matrix to the FMN
and, as a result, FMNH2is formed. At this time, a pair of protons belonging to NAD
is pushed into the intermembrane space.
The structure of the mitochondria
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