AL Bio Notes Respiration P.1

AL Bio notes Respiration P.1

Respiration

O

rganisms maintain themselves by the continual expenditure of energy. Most cells derive this energy from the breakdown of food molecules, a process known as cellular or internal respiration. Substances that are broken down to yield energy are called respiratory substrates.

In most organisms glucose is broken down to carbon dioxide and water in a process which requires molecules of oxygen, and is known asaerobic respiration.

C6H12O6 + 602 6C02 + 6H2O + energy

glucose oxygen carbon dioxide water

In the absence of oxygen molecules, anaerobic respiration takes place in which glucose is partiallybroken down, to lactic acid in animals, and to carbon dioxide and alcohol in plants. The latter process is also called alcoholic fermentation:

anaerobic respiration in plants C6H12O6 C2H5OH + 2CO2 + C02 + energy (ethyl alcohol)

anaerobic respiration in animals C6H12O6 CH3CHOH.COOH + energy

(lactic acid)

The partial breakdown of glucose in anaerobic respiration yields only a fraction of the chemical energy stored in the molecule.

Glucose is broken down in a series of carefully controlled reactions, during which energyis made available in relatively small amounts which can be harnessed and made to perform work.
A much smaller proportion of the energy in the glucose molecule is dissipated as heat compared with combustion. (at most about forty per cent). /

The energy made available during the respiratory process is stored in molecules ofadenosine triphosphate, usually called ATP.

AL Bio notes Respiration P.1

I)THE ROLE OF ATP AS A STORE AND SUPPLIER OF ENERGY

ATP occurs in all living cells. It is the un______energy 'currency'. All energy-spending activities such as synthesis, muscle contraction, transmission of nerve impulses …….., depend on ATP as theirim______ source of energy.
Energy is supplied by the splitting of the bond that attaches the terminal phosphate groups to the rest of the molecule. The bond is broken by hydrolysis. This reaction yields a large amount of energy (about 33 kJ mol-1). Because of this high energy yield, ATP is often called an 'energy-rich' or 'high-energy' molecule.
ATP thus provides a source of energy that is easily and rapidly accessible to any energy-requiring process. /
Matching : Analogy of Energy availability and Money currency
ATP /  /  / Investment accounts
glucose /  /  / deposit accounts
carbohydrate reserves, starch and glycogen /  /  / cash
fat reserves /  /  / cheque

II)THE FORMATION OF ATP

Molecules of ATP are constantly being 'spent' and must be replaced. They are formed from ADP and inorganic phosphate by condensation. This reaction is also aphosphorylation.
The regeneration of ATP requires a supply of energy = that which it gives on hydrolysis. This is supplied by respiration.
There are two mechanisms that direct the energy yield from respiration into the formation of ATP; namely s______-linked phosphorylation and o______phosphorylation.

a)Substrate-linked phosphorylation

At 3 steps in the respiratory sequence where p______is detached from one of the intermediate compounds of glucose breakdown (the respiratory substrate), together with sufficient energy to form a bond with ADP. This process is thus substrate-based or linked and, unlike oxidative phosphorylation, does notneed O2.

b)Oxidative phosphorylation

This process involves oxidation reactions. Such reactions tend to have a high ‘energy yield’, and it is this energy that is used to combine inorganic phosphate and ADP. /

The following types of reaction are all described as Oxidations:

i)The addition of oxygen to a molecule,

e.g. C + O2 CO2 (the burning of carbon in air)

ii)The removal of hydrogen from a molecule (dehydrogenation).

e.g. AH2 + B A + BH2

iii)The removal of an electron from a charged ion,

e.g. Fe ++ - e (an electron) Fe+++

All three types of oxidation are involved in oxidative phosphorylation :

1)Pairs of hydrogen atoms are removed from a number of respiratory intermediates by dehydrogenase enzymes; the respiratory intermediates are thus oxidised.

2)The hydrogen removed is transferred tohydrogen-acceptors, usually nicotinamide adenine dinucleotide (NAD+). These molecules are also described as coenzymesbecause they are essential for the functioning of the dehydrogenase enzymes.

3)The hydrogen is split into protons and electrons on its way through the series of carriers, the electrons are eventually combined into with oxygen to form water. The whole sequence of carriers, including the initial hydrogen-acceptors, comprisesthe Electron transport chain or Respiratory chain.

c)Electron Transport Chain (ETC) / The Respiratory Chain

A series of coupled ox______/re______reactions where e______are passed like hot potatoes from one membrane-bound p______to another before being finally attached to a terminal electron acceptor (o______in respiration, N______in photosynthesis ). ATP is formed inthe process.

Each of the carriers in the respiratory chain is successively reduced and then oxidised as it first accepts and then hands on the hydrogen. ETS moves both electrons and protons: electrons are passed from carrier to carrier in the membrane, while protons are moved from in_____ to out____ of membrane (inner membrane).

In fact, early in the series of protein carriers, it is only the electrons of the hydrogen atoms that are passed along the chain. Hydrogen atoms are split into hydrogen ions and electrons.

H H + + e- (an electron)

Each carrier in this chain has a greater affinity for electrons than its predecessor so there is a one-way flow of electrons along the chain. In particular the electron transport chain contains severalcytochromes, which are a group of iron-containing proteins (h____group). The state of the iron is altered from the oxidised ferric (Fe + + +) form to the reduced ferrous (Fe ++) form, and then back to the oxidised form as the cytochrome accepts and then passes on electrons.

At the final cytochromes in the chain, the electrons combine with hydrogen ions, and then combine with molecules of oxygen to form water.

2H+ + 2e- 2H

2H+ + O2 H2 O

Incidentally, cyanide exerts its toxic effects bybinding cytochrome oxidase so as to prevent the binding of o______. It thus acts as a respiratory in______, and in large amounts can rapidly cause death through energy lack.

The energy yield is on average 3ATP from each NADH entering the ETC.

Reference reading: Proton gradient / Chemiosmotic hypothesis (Peter Mitchell, 1961)
As electrons flow through ETC, at certain steps protons (H+) are moved from inside to outside of the membrane. This builds up proton gradient; since + charges are removed from inside of cell, -ve charge remains inside, mainly as OH- ions. pH just outside membrane can reach 5.5, pH just inside membrane can reach 8.5 ---> difference of 3 pH units, or 1000x concentration differential of H+ across membrane. This represents potential energy stored up in proton gradient.

1. Protons are translocated across the membrane, from the matrix to the intermembrane space
2. Electrons are transported along the membrane, through a series of protein carriers
3. Oxygen is the terminal electron acceptor, combining with electrons and H+ ions to produce water
4. As NADH delivers more H+ and electrons into the ETS, the proton gradient increases, with H+ building up outside the inner mitochondrial membrane, and OH- inside the membrane.

Membrane is basically impermeable to protons, so gradient doesn't get wasted away by leaky reentry.
ATP synthase protein complex contains onlychannels for proton entry.
As protons push in through channel : ADP + Pi ---> ATP.
This can be called chemiosmotic phosphorylation or oxidative phosphorylation.

III)THE BREAKDOWN OF GLUCOSE

Aerobic and anaerobic respiration share a common initial pathway known as glycolysis (which means sugar-splitting), in which the glucose is broken down through a series of reactions to pyruvic acid (2-oxopropanoic). The breakdown of glucose to pyruvic acid takes place in the c______and does not require the presence of oxygen molecules.

An outline of glycolysis

This is the initial pathway shared by both aerobic and anaerobic respiration. The process takes place in the cytoplasm and is not dependent on the presence of oxygen.

It comprises two stages; in the first, glucose is phosphorylated to fructose disphosphate. After phosphorylation, the substrates were eventually broken down to two pyruvic acid molecules.

Each of the reactions is catalysed by an enzyme.

Glucose

2ATP

2ADP

Fructose diphosphate

Glyceraldehyde Dihydroxyacetone

phosphate phosphate

NAD+

NADH &

H+

inorganic

phosphate

Diphosphoglyceric acid

ADP

ATP

Phosphoglyceric acid

ADP

ATP

a)The Energy yield of Glycolysis

AL Bio notes Respiration P.1

Summary of glycolysis
input / output
One glucose molecule / Two pyruvic acid molecules
2ATP
(for phosphorylation of glucose and fructose) / 4ATP from substrate level phosphorylation
2ADP and Pi
2 NAD+ / (NADH & H+ ) X 2

Thus, the net yield is two molecules of ATP per glucose molecule. However, if oxygen is present the two pairs of hydrogen atoms removed by NAD+ (one pair from each triose phosphate) can be passed to the ETC andwill there yield a further six molecules of ATP (by oxidative phosphorylation).

Thus in conditions where oxygen is available the net gain from glycolysis is eight molecules of ATP per glucose molecule.

AL Bio notes Respiration P.1

The fate of pyruvic acid differs according to whether oxygen is present or not and, in the latter circumstance, according to whether plant or animal cells are being considered.

/ ii)The final anaerobic pathway in animals
In animal cells the pyruvic acid is reduced by the NADH, formed during glycolysis, to lactic (2-hydroxypropanoic) acid.
This alsoregenerates the NAD+ (coenzyme for dehydrogenation) needed in glycolysis.
i)The final anaerobic pathway in plants
In the absence of oxygen pyruvic acid is converted to acetaldehyde (ethana1) and carbon dioxide.
The NADHformed during glycolysis is then used to reduce the acetaldehyde to ethanol.
The oxidised NAD+ is thenregenerated to pick up more hydrogen atoms from glycolysis.

b)The Energy yield of Anaerobic respiration

The net yield from anaerobic respiration is thus 2 ATP per glucose molecule. Two moles of ATP store about 66kJ. This is a small return from glucose, which stores about 2800 kJmol -1.

IV)THE AEROBIC PATHWAY

If oxygen is available pyruvic acid enters mitochondria and there embarks on a series of reactions that involves the removal of Hatoms to the ETC--and the removal of CO2 molecules. The removal of CO2 isalways linked to a dehydrogenation and so is called oxidative decarboxylation.

Before entering the cycle, pyruvic acid (3C) is converted to acetyl coenzyme A (2C)(acetyl-CoA) by oxidative decarboxylation. The Krebs cycle may be summarized into 4 major steps :

1.Formation of a 6-C citric acid molecule

Acetyl-CoA (2C) combines with oxaloacetic acid (4C), to form citric acid (6C).

2.Oxidation of the 6-C molecule

Citric acid (6C) is oxidized to -Ketoglutaric acid (5C) by oxidative decarboxylation.

1. Oxidation of the 5-C molecule

-Ketoglutaric acid is oxidized, to a 4-C intermediate by oxidative decarboxylation.

4.Oxidation and regeneration of Oxaloacetic acid

Further dehydrogenation occurs and oxaloacetic acid is regenerated. The cycle starts again with another acetyl-CoA produced from pyruvic acid.

/ The Krebs or Tricarboxylic acid cycle (TCA cycle)
Take place in the matrix of mitochondria.
only operate if oxygen is available to act as the final electron acceptor.
6-carbon citric acid is gradually broken down by four dehydrogenations and two decarboxylations to reform the 4-carbon oxaloacetic acid.
Taking into account the oxidative decarboxylation when pyruvic acid are converted into acetyl CoA, three CO2 molecules and 5 pairs of Hydrogens are removed.

a)The Energy yield of Aerobic respiration

5 pairs of H atoms are removed as pyruvic acid is broken down in the aerobic pathway, 4 pairs from each turn of the Krebs cycle and one pair from the initial conversion of pyruvic acid into acetyl-CoA. The total yield from this part of the respiratory process is thus ten pairs of hydrogen atoms per glucose molecule (each glucose molecule is broken down to two molecules of pyruvic acid).

8 pairs are removed by NAD+, each pair providing sufficient energy to form 3 ATP molecules. The 2 pairs removed by FAD each provides enough energy to form 2 ATP molecules. To these must be added the 2 pairs of hydrogen atoms removed during glycolysis. These are passed via a carrier/ shuttle in the mitochondrial membrane to either NAD+ or FAD within the mitochondria (varies according to tissue type) resulting in the formation of either 6 or 4 ATP molecules. The total number of ATP molecules formed by oxidative phosphorylation is thus :

(8 x 3) + (2 x 2) + 6 or 4 = 32 or 34.

To this must be added the 4 ATP molecules formed by substrate-linked phosphorylation ( 2 in glycolysis, 2 in krebs cycle). The total yield from aerobic respiration is thus 36 or 38 molecules of ATP per glucose molecule.

ATP yield during aerobic respiration of one molecule of glucose
Respiratory process / No.of reduced hydrogen carrier molecules formed / No. of ATP molecules formed from ETC / No. of ATP molecules from substrate-level phosphorylation / Total No.of ATP molecules
glucose to pyruvate (Glycolysis)
pyruvate to acetyl CoA
Krebs (TCA) cycle
Total ATP = / 38

Q.Below is called Warburg apparatus which has been designed to measure the effect of various substances on the rate of respiration. Mung beans seeds were crushed with isotonic buffer enriched with glucose in cold. Its extract was made by a high speed differential centrifugation. The supernatant made by 1,000 g centrifugation was obtained and used in this apparatus.

a)Why was it necessary to crush the seeds with isotonic buffer enriched with glucose in cold ?(4m)

b)What essential component of the cell must be present in the extract? Why was it necessary?(2m)

c)Why should the KOH be separated from the mung bean extract? What was the purpose of using the filter paper? (2m)

d)After setting up the apparatus, its was left undisturbed. The manometric reading was recorded at one minute intervals. At ten minutes, the Warburg apparatus was tilted slightly to mix the ADP from the side arm to the mung bean extract. The result was recorded by graph I below.

i)Explain the shape of graph I

I)before 10 minutes.(2m)

II)after 10 minutes.(2m)

ii)Identify two possible sources of errors which leaded to inaccuracy of the experiment. Suggest two ways in which these errors could be minimized. (4m)

e)Another experiment was performed to demonstrate the effect of substance X on respiration. Substance X was placed in the side arm of the Warburg apparatus instead of ADP. The same experiment was performed and substance X was added to the extract at 20 minutes after commencement of the experiment. The result was recorded in graph II above. Name a possible substance for 'X', explain how it might lead to the effect as shown in graph II. (4m)

b)The Site of the Aerobic process

Mitochondria tend to be most abundant in cells with a high e______requirement.

They are sausage-shaped organelles comprising two m______layers. Within the inner membrane encloses an aqueous solution, the m______. The inner membrane is folded into c______ which project into the matrix.

The matrix contains the en______ involved in the Krebs cycle and the breakdown of fatty acids, while theinner mitochondrial membrane contains the en and electron c of the electron transport chain (ETC).

There are tiny spheres on the inner membrane which are believed be the sites of ATP synthase, the enzyme that brings about the formation of _____ from ADP and inorganic phosphate.. The cristae presumably serve to increase the surface area for enzyme and carrier attachment.

Supporting this, it has been observed that the cristae are more numerous in cells with a high energy requirement.

Draw an annotated diagram to show how the structure of a mitochondrion is related to its function (5m)
/

V)RESPIRATION USING OTHER SUBSTRATES

Carbohydrates are not the only source of energy.
In certain circumstances, fats and proteins can supply part of the organism's energy needs.
The breakdown of fats and proteins is largely dependent on the presence of o______ as they are converted into substances, most of which enter the Krebs cycle where they are oxidised to yield their energy. /

a)The use of fats as a respiratory substrate

Fats form the long-term energy stores in many organisms. They are generally used only when the carbohydrate reserves are exhausted. Fats usually have amuch higher energy yield than carbohydrate. Some tissues like liver, and seeds possessing large deposits of fats and are able to use fat directly as a respiratory substrate without first converting it to carbohydrate.

The fats are first removed from the fat depots (under the skin and around various internal organs), a process known as mobilisation. They are then transported to the liver where they split into fatty acids and glycerol.

The glycerol is phosphorylated to form triose phosphate / glyceraldehydes phosphate which enters the glycolytic sequence.

Fatty acid molecules are oxidized by a process called β-oxidation, which involves 2C fragments of acetyl CoA being split off from the acid until the entire fatty acid molecule has been broken up and oxidised. This takes place in the matrix of the mitochondria.

Each acetyl CoA formed can enter the Krebs cycle. The longer the fatty acid chain, the greater will be the energy yield. E.g 147 ATP can be obtained from one stearic acid molecules (C17H35COOH).

b)Protein as a respiratory substrate

Many plant seeds employ protein as an energy storage material. Animals do not generally employ protein as an energy reserve; they do so at the expense of their own tissue proteins. Accordingly, proteins are only respired by animals as a last resort in cases of prolonged starvation.

When proteins are respired they are first broken down into amino acids. These are then deaminated, that is, their nitrogen-containing amino group is removed. The products of deamination are ammonia and an α-keto acid

Krebs cycle