In this article some important principles are undertaken so that we can well understand the various mechanisms of energy conservation.
In microbiology, energy is measured in units of kilojoules (kJ), a measure of heat energy. Chemical reactions are accompanied by changes in energy. Although in any chemical reaction some energy is lost as heat, in microbiology the interest is in free energy (abbreviated G), which is defined as the energy released that is available to do useful work.
The change in free energy during a reaction is expressed as ∆G0,, where the symbol A should be read “change in”. The “o’ and ” ‘ “(prime) mean that the free energy value was obtained under “standard’ conditions: pH 7, 25°C, all reactants and products initially at 1M concentration.
If in the reaction;
A + B = C + D
the ∆G0, is negative, the reaction will proceed with the release of free energy, energy that the cell may be able to conserve in the form of adinosine triphosphate (ATP). Such energy yielding reactions are called exergonic. However, if ∆G0, is positive the reaction requires energy in order to proceed, such reactions are called endergonic. Thus, from the standpoint of the microbial cell, exergonic reactions yield energy, while endergonic reactions require energy.
Free Energy of Formation:
Free energy of formation (abbreviated GOF) is the energy yielded or energy required for the formation of a given molecule from its constituent elements. By convention, the free energy of formation (GOF) of the elements (for example, C, H2, N2) is zero.
If the formation of a compound from elements proceeds exergonically, then the free energy of formation of the compound is negative (energy is released), whereas if the reaction is endergonic (energy is required), then the free energy of formation of the compound is positive.
The values of free energy of formation are always in kilojules/molecule (kJ/mol). Using free energies of formation, it is possible to calculate the change in the free energy taking place in a given reaction. For a simple reaction such as A + B → C + D. the change in free energy (∆G0,) can be calculated by subtracting the sum of the free energies of formation of the reactants (in this case A and B) from that of the products (in this case C and D).
Change in free energy (∆G0, ) of
A + B → C + D = GOF [C + D] – GOF [ A + B]
Table 23.1 shows the free energies of formation for some of the compounds of biological interest. For most of the compounds taken in this table the free energy of formation (ΔGOF) is negative, reflecting the fact that compounds tend to form spontaneously from elements. However, the positive free energy of formation for nitrous oxide (N2O; + 104.2 kJ/mol) tells us that this molecule does not form spontaneously, but rather decomposes to nitrogen and oxygen.
Oxidation-Reduction (Redox) Reactions:
The free energy in living organisms is conserved involving oxidation-reduction (redox) reactions. Oxidation- reduction (redox) reactions are those in which electrons are donated by an electron donor (oxidation) and are accepted by an electron acceptor (reduction).
The electron donor is called the reducing agent or reductant, whereas the electron acceptor as oxidising agent or oxidant. By convention, such a reaction is written with the reductant to the right of the oxidant and the number (n) of electrons (e–) donated.
Oxidant + ne– ⇋ reductant
Electron Donors and Electron Acceptors:
Oxidation-reduction reactions, as stated earlier, involve electrons being donated by an electron donor and being accepted by an electron acceptor. However, the electrons released by the electron donor cannot exist free in solution; they must be subsequently accepted by an electron acceptor and become the part of it. This is the reason why for any oxidation to occur, a subsequent reduction must also occur.
For example, hydrogen gas (H2) can release electrons and hydrogen ions (protons) and become oxidized:
H2 →2e- + 2H+
The above reaction is only a half reaction and needs subsequently the second half reaction to complete.
In second half reaction there can be the reduction of many different substances including O2:
½ 02 + 2e– + 2H+ →H2O
The second half reaction (reduction reaction), when coupled to the first half reaction (oxidation reaction), yields the following overall balanced reaction:
H2 + ½ O2 →H2O
In the above overall balanced reaction, one refers to the H2 oxidised (i.e., electron donor) and O2 reduced (i.e., electron acceptor). The overall view of the formation of H2O from the electron donor H2 and the electron acceptor O2 is shown in Fig. 23.1.
Reduction potentials (E0’) is the expression of the tendency of substances to become oxidised or to become reduced; the substances vary in the tendency to become oxidised or to become reduced.
The reduction potential is measured electrically in units of volts or millivolts in reference to a standard substance; the reduction potential of hydrogen (H2) at pH 7 is -0.42 volts or -0.420 millivolts. pH 7 is used because it refers to neutrality and the cytoplasm of most cells is neutral or nearly so.
Most molecules can be either electron donors or electrons acceptors under different conditions, depending on the substances with which they react. The same atom on each side of the arrow in the half reactions can be thought of as representing a redox couple. When writing a redox couple, the oxidised form is always placed on the left.
In constructing complete oxidation-reduction reactions from their constituent half reactions, it is simplest to remember that the reduced substance of a redox couple whose reduction potential is more negative donates electrons to the oxidized substance of a redox couple whose reduction potential is more positive.
Thus, in a redox couple 2H+ / H2, which has a reduction potential of -0.42 volts, H2has a great tendency to donate electrons. On the other hand, in the redox couple ½ O2/H2O, which has a potential of +0.82 volts, H2O has a very slight tendency to donate electrons, but O2 has a great tendency to accept electrons.
It follows then that in a reaction of H2 and O2, H2 will be the electron donor and become oxidised, and O2 will be the electron acceptor and become reduced (Fig. 23.1).
Electron toner is an imaginary vertical tower that represents the range of reduction potentials for redox couples from the most negative at the top to the most positive at the bottom (Fig. 23.2).
The reduced substance in the redox pair at the top of the tower possesses the greatest tendency to donate electrons, whereas the oxidised substance in the couple at the bottom of the tower has the greatest tendency to accept electrons. As electrons from the electron donor at the top of the tower fall, they can be “caught” by acceptors at various levels of the tower.
The farther the electrons drop from a donor before they are “caught” by an acceptor, the greater the amount of energy released. O2, at the bottom of the tower, is the most favourable electron acceptor used by organisms. In the middle of the electron tower, redox couples can act as either electron donors or electron acceptors.
For instance, under conditions where oxygen is absent (called anoxic) in the presence of H2, fumarate can be electron acceptor (producing succinate), and under other conditions where oxygen is present (called aerobic) in the absence of H2, succinate can be an electron donor (producing fumarate).
The transfer of electrons in an oxidation-reduction reaction from donor to acceptor in a cell normally involves one or more intermediates that are called electron carriers (or carriers). In such conditions the initial electron donor is called primary electron donor, whereas the final electron acceptor as the terminal electron acceptor.
The net change in the free energy (∆G0,) of the complete reaction sequence is determined by the “difference” in the reduction potentials (E0’) between the primary electron donor and the terminal electron acceptor.
Electron carriers can be divided into two general groups:
(1) Freely diffusible and (2) firmly attached (fixed) to enzymes in the cytoplasmic membrane. Freely diffusible carriers include the coenzymes nicotinamide adenine dinucleotide (NAD+) and NAD-phosphate (NADP+) , whereas membrane-associated electron carriers include NADH dehydrogenases, flavoproteins that contain flavin mononucleotide (FMN) or flavin-adenine dinucleotide (FAD), cytochromes, nonheme iron-sulphur (Fe/S) proteins (ferrodoxin) and quinones.
Nicotinamide Adenine Dinucleotide (NAD+) and NAD-phosphate (NADP+):
These are the coenzymes that act as freely diffusible electron carriers and transport electrons between two different locations. The nicotinamide ring of NAD+ and NADP+ (Fig. 23.3) accepts two electrons and one proton from a donor, while a second proton is released. The reduction potential of the redox couple NAD+/NADH (or NADP+/NADPH) is -0.32 volt, which places it fairly high on the electron tower, i.e., NADH (or NADPH) is a good electron donor.
However, although the NAD and NADP+ couples possess the same reduction potentials, they generally function in different capacities in the cell. NAD+/NADH is directly involved in energy generating (catabolic) reactions, whereas NADP+/NADPH is involved primarily in biosynthetic (anabolic) reactions.
NADH dehydrogenases are proteins bound to the inside surface of the cell membrane. They accept hydrogen atoms from NADH generated in various cellular reactions and pass the hydrogen atoms to flavoproteins.
Flavoproteins are proteins possessing a derivative of riboflavin. Flavoproteins accept hydrogen atoms and donate electrons. Two flavoproteins are commonly found in cells—flavin mononucleotide (FMN) and flavin- adenine dinucleotide (FAD) (Fig. 23.4).
Flavin mononucleotide (FMN) is bonded to ribose and adenine through a second phosphate. However, these two flavoproteins bear two electrons and two protons (two hydrogen atoms) on their complex ring system. Riboflavin, also called vitamin B2, is a required growth factor for some organisms.
Cytochromes are proteins with iron-containing porphyrin ring (Fig. 23.5) also called heme. The cytochromes undergo oxidation and reduction through loss or gain of a single electron by the iron atom centrally placed in the porphyrin ring of the cytochrome:
Cytochrome – Fe2+ ⇋ Cytochrome – Fe3+ + e–
Cytochromes do not carry hydrogen atoms (protons). Several different cytochromes (cyt b, cyt c, etc.) are a prominent part of respiratory electron transport chains.
Nonheme Iron-Sulphur (Fe/S) Proteins:
Some iron-sulphur (Fe/S) containing electron carrying membrane-associated proteins lack a heme group and are called nonheme iron-sulphur (Fe/S) proteins. Various arrangements of iron and sulphur have been found in different nonheme iron-sulphur proteins, but Fe2S2 and Fe4S4 clusters are the most common.
The iron atoms are bonded to free sulphur and to the protein via sulphur atoms from cysteine residues (Fig. 23.6). Ferredoxin is a common iron-sulphur protein of Fe2S2 configuration occurring in biological systems.
This electron carrier is active in photosynthetic electron transport and several other electron transport processes. Since the reduction potentials of iron-sulphur proteins vary over a wide range depending on the number of iron and sulphur atoms and their attachment-pattern to protein, different iron-sulphur proteins function at different points in an electron transport process. Like cytochromes, these proteins also carry electrons only, not hydrogen atoms.
Quinones are membrane-associated highly hydrophobic non-protein-containing molecules that act as electron carriers in electron transport processes. Some quinones occurring in bacteria are related to vitamin K, a growth factor for higher animals.
Like flavoproteins, quinones serve as proton (hydrogen atom) acceptors and electron donors. Coenzyme Q (CoQ) or ubiquinone is a quinone that carries electrons and protons (hydrogen atoms) in many respiratory electron transport processes (Fig. 23.7).
Energy released as a result of oxidation-reduction reactions must be conserved so that it can be utilized wherever and whenever required in the cellular functions. Conservation of energy in living organisms is made in high-energy phosphate bonds of high-energy phosphate compounds (e.g., phosphoenolpyruvate, 1,3- bisphosphaglycerate, ATP, ADP, etc.) and thio-ester bonds of the derivatives of coenzyme A (e.g., acetyl- CoA). We take only ATP and coenzyme A (CoA) derivatives (e.g., acetyl-CoA) for further consideration here.
Adenosine Triphosphate (ATP):
The most important high-energy phosphate compound in living organisms is adenosine triphosphate (ATP), a practical form of major energy-currency the cells possess to carry out their work. ATP consists of the ribonucleoside adenosine to which three phosphate molecules are bonded in series (Fig. 23.8).
Out of the three phosphate bonds of ATP, as is apparent in the Fig. 23.8, two are high-energy anhydride bonds having high free energies of hydrolysis, whereas one is low-energy ester bond.
When ATP breaks down to adenosine diphosphate (ADP) and orthrophosphate (Pi) as a result of the hydrolysis of high energy anhydride bond, the free energy is made available to drive biosynthetic reactions and other aspects of cell function through carefully regulated processes in which the energy released from ATP hydrolysis is coupled to energy-requiring reactions. Later, energy from photosynthesis, aerobic respiration, anaerobic respiration, and fermentation is used to resynthesise ATP from ADP and Pi.
Coenzyme A (CoA) Derivatives (Acetyl-CoA):
Coenzyme A (CoA) derivatives are certain other high-energy compounds that are produced in the cell and can conserve the energy released in oxidation-reduction reactions. These derivatives [acetyl-CoA (Fig. 23.9) is just one of many CoA derivatives] possess thio-ester (sulphoanhydride) bonds instead of phosphoanhydride bonds that occur in high energy phosphate compounds (e.g., ATP). CoA derivatives yield sufficient free energy on hydrolysis, which is used to drive the synthesis of a high energy phosphate bond in energy metabolism and biosynthesis of fatty acids.
For instance, in the reaction:
Acetyl-CoA + H2O + ADP + P → Acetate– + HS-CoA + ATP + H+
the energy released in the hydrolysis of coenzyme A is conserved in the synthesis of ATP. CoA derivatives play very important part in the energy conservation of anaerobic microorganisms, especially in those whose energy metabolism involves fermentation.
Options for Energy Conservation:
Metabolism is the sum total of all biochemical reactions that take place in the cell with the involvement of flow of energy and the participation of variety of enzymes and proteins. Metabolism, in fact, represents the chemistry of life and can be divided into two major parts: catabolism and anabolism.
Catabolism (Gk. cata = down, ballein = to through) represents the breakdown of more complex chemicals into smaller, simpler molecules resulting in the release of energy. Some part of this released energy is trapped and made available for cellular functions while the rest is released as heat. In anabolism (Gk. ana = up, ballein = to through) the similar molecules are used in the synthesis of complex molecules with energy utilization.
Enzymes required for metabolic activities are synthesized in the cell, whereas energy is obtained from one of the three sources (Fig. 23.10):
(i) Chemolithotrophic microbes carry out oxidation of inorganic chemicals that releases energy,
(ii) chemoorganotrophic microorganisms oxidize organic molecules to liberate energy, and
(iii) phototrophic microorganisms trap radiant energy of sun by the process of photosynthesis.
The chemotrophic macroorganisms (both chemolithotrophic and chemoorganotrophic), the microorganisms that use chemicals as electron donors in their energy metabolism, have adopted two catabolic mechanisms for energy conservation respiration and fermentation. In respiration, the energy is conserved by the process oxydative phosphorylation with the involvement of molecular oxygen or some other externally derived electron-acceptor.
Respiration, however, is of two different types, namely, aerobic and anacrobic. In aerobic respiration, the final electron acceptor is oxygen whereas the electron-acceptor in anaerobic respiration is more often inorganic (e.g., NO3–, SO42-, CO2, Fe3+, SeO42-, and many others), though organic electron-acceptors such as fumaric acid may also be used.
In fermentation, the energy is produced by substrate-level-phosphorylation in which ATP is synthesized as a result of the oxidation of an organic compound without involvement of any usable external electron-acceptor. Phototrophic microorganisms employ anabolic mechanism and trap light energy of sun during photosynthesis (synthesis of complex molecule using simpler molecules) by the process photophosphorylation.