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Introduction
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The sum total of all the chemical conversions in a cell is called metabolism.
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The chemical buildup and breakdown of substances requires energy transformations mediated by enzymes.
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In some cases, enzymes represent valuable targets for therapeutic drugs. Blocking the activity of the COX-2
enzyme, for example, helps to minimize inflammation in the joints.
Energy and Energy Conversions
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To physicists, energy represents the capacity to do work.
To biochemists, energy represents the capacity for change.
Cells must acquire energy from their environment.
Cells cannot make energy; energy is neither created nor destroyed, but can be transformed.
In life, energy transformations consist primarily of molecular movement and changes in chemical bonds.
Energy changes are related to changes in matter
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There are two main types of energy: kinetic energy and potential energy. (See Figure 6.1.)
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Potential energy is energy of state or position—it is stored energy. Potential energy is like money in the bank;
it remains there until you spend it.
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Potential energy is “spent” in the form of kinetic energy, which is energy in action. Kinetic energy does work
that alters the state or motion of matter.
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Metabolism can be divided into two types of activities: anabolic reactions and catabolic reactions.
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Anabolic reactions link simple molecules together to make complex ones. These are energy-storing reactions.
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Catabolic reactions break down complex molecules into simpler ones. Some of these reactions provide the
energy for anabolic reactions.
Thermodynamics
The first law: Energy is neither created nor destroyed
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Life follows the laws of physics, including the laws of thermodynamics, which apply to the whole universe or
to any closed system within the universe.
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Although living cells are open systems (they exchange matter and energy with their surroundings), they still
obey these laws.
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First law of thermodynamics: During any conversion of forms of energy, the total initial energy will equal the
total final energy. (See Figure 6.2a.)
The second law: Not all energy can be used, and disorder tends to increase
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Second law of thermodynamics: When energy is transformed, some is unavailable to do work. (See Figure
6.2b.)
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Theoretically, a process could be 100 percent efficient, but efficiency certainly could not be greater than 100
percent.
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Energy is never utilized perfectly in a physical process or chemical reaction, so the efficiency is less than 100
percent.
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Total energy (enthalphy, or H) = usable energy (free energy, or G) + unusable energy (absolute temperature,
T  entropy, S): H = G + TS.
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Usable energy: G = H – TS.
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Although G, H, and S cannot be measured precisely, change at a constant temperature can be measured
precisely and calculated in calories or joules.
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The equation G = H – TS (with delta representing the change in a value) describes the events in terms of
changes in energy that occur during chemical reactions.
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Notice that there is no  in front of T; T must be constant in order for other variables to be calculated.
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If G is positive (+), then the reaction requires an input of energy. This is the case for anabolic reactions.
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If G is negative (–), energy is released. This is the case for catabolic reactions.
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The efficiency of energy use is dependent on the system. If two different automobiles weighing the same
amount require different amounts of fuel to go the same distance, the one that uses less fuel is more efficient.
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Some biological systems are amazingly efficient, although never 100 percent so. An example of an efficient
use of free energy in living cells is the generation of ATP from glucose, when oxygen is used as the final electron
acceptor (see Chapter 7). Although almost two dozen enzyme-catalyzed reactions take place, the net capture of energy
is about 68 percent!
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If a chemical reaction increases entropy, its products are more disordered or random than its reactants are. An
example of such a reaction is the hydrolysis of a protein to its amino acids.
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When amino acids are freed from proteins, energy is released, G is negative, and although the released
energy usually is not captured in a useful form for the cell, there is an increase in entropy and a release of heat.
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When proteins are made from amino acids, energy is used, there are fewer products, and S is negative.
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The second law of thermodynamics also predicts that, as a result of energy conversions, disorder tends to
increase.
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This tendency for disorder to increase gives a directionality to physical and chemical processes, explaining
why some reactions proceed in one direction rather than another. (See Figure 6.2b.)
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It may seem that highly complex organisms, such as the human body, are in apparent disagreement with the
second law, but this is not the case.
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The metabolic processes that take place in living tissues produce far more disorder than the order present
within the tissues.
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To maintain order, life requires a constant input of energy.
Chemical reactions release or take up energy
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Anabolic reactions may make single products from many smaller units; such reactions consume energy.
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Catabolic reactions may reduce an organized substance (e.g., a glucose molecule) into smaller, more
randomly distributed substances (e.g., carbon dioxide and water). Such reactions release energy.
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There is a direct relationship between the amount of energy released by a reaction (–G), or the amount taken
up (+G), and the tendency of a reaction to run to completion without an input of energy.
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A spontaneous reaction goes more than halfway to completion without input of energy, whereas a
nonspontaneous reaction proceeds that far only with an input of energy. (See Figures 6.3a and 6.3b.)
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Spontaneous reactions are called exergonic and have negative G values (they release energy).
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Nonspontaneous reactions are called endergonic and have positive G values (they consume energy).
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If under certain conditions A  B is spontaneous (and exergonic), then B  A must be nonspontaneous (and
endergonic).
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Making protein is endergonic; hydrolyzing protein is exergonic.
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In principle, all reactions are reversible (A  B).
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Adding more A speeds up the forward reaction, A  B; adding more B speeds up the reverse reaction, B 
A.
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At the point of chemical equilibrium, the relative concentrations of A and B are such that forward and reverse
reactions take place at the same rate. (See Figure 6.4.)
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Although no further net change occurs at this point, reactions of individual molecules continue.
Chemical equilibrium and free energy are related
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An example of equilibrium can be seen in the cellular conversion of glucose 1-phosphate to glucose 6phosphate.
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At pH 7 and 25C, the concentration of the product rises while the concentration of the reactant falls.
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Equilibrium is reached when the product-to-reactant ratio is 19:1.
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At this point the forward reaction has gone 95 percent to completion.
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The further a reaction goes toward completion in order to reach equilibrium, the greater the amount of free
energy released.
ATP: Transferring Energy in Cells
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All living cells use adenosine triphosphate (ATP) for capture, transfer, and storage of energy. Figure 6.5
shows the chemical diagram of ATP.
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ATP may be thought of as the energy “currency” of the cell; some of the free energy released by certain
exergonic reactions is captured in ATP, which then can release free energy to drive endergonic reactions.
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ATP is not an unusual molecule and it has other uses as well; for example, it can be converted into a building
block for DNA and RNA.
ATP hydrolysis releases energy
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ATP consists of the nitrogenous base adenine bonded to ribose. Carbon 5 of the ribose has three phosphate
groups attached.
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ATP can hydrolyze to yield ADP and an inorganic phosphate ion (P i short for HPO42).
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ATP + H2O  ADP + Pi + free energy. The change in free energy (G) is –12 kcal/mol at a living cell’s
typical temperature and pH.
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The equilibrium is far to the right of the equation, toward ADP production; there are 10  106 ADP
molecules to each remaining ATP.
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Making ATP from ADP involves overcoming repulsive negative charges on the phosphates to be joined. The
energy to do this is stored in glucose or other fuel molecules and released in the endergonic process. (See Figure 6.6.)
ATP couples exergonic and endergonic reactions
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The reverse reaction of ATP hydrolysis, the formation of ATP from ADP and P i, is endergonic and consumes
as much free energy as is released by the breakdown of ATP: ADP + P i + free energy  ATP + H2O.
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Many different exergonic reactions in the cell can provide the energy to convert ADP to ATP; in eukaryotic
cellular respiration, the energy released from fuel molecules such as glucose is captured in ATP.
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ATP shuttles energy from exergonic reactions to endergonic reactions. Figure 6.7 illustrates a coupled
reaction that uses ATP to provide energy for the synthesis of glutamine from an ammonium ion and glutamate.
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Each cell requires millions of molecules of ATP per second to drive its biochemical machinery.
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Each ATP molecule undergoes about 10,000 cycles of synthesis and hydrolysis every day.
Enzymes: Biological Catalysts
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A catalyst is any substance that speeds up a chemical reaction without itself being used up. Living cells use
biological catalysts to increase rates of chemical reactions.
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Most biological catalysts are proteins called enzymes. Certain RNA molecules called ribozymes also are
catalysts.
For a reaction to proceed, an energy barrier must be overcome
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It is easy to predict the direction of a spontaneous reaction, but not the likelihood or rate of the reaction.
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For example, knowing the direction of the spontaneous reaction of wood with oxygen and the levels of endproducts in our environment allows us to predict that forests will burn. However, it might be a few years or a few
hundred years before this burning occurs.
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A forest fire is initiated by an initial investment of energy (e.g., from a lightning strike), and then is
perpetuated by energy released from the combustion of wood, which is invested in the unburned molecules.
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The energy that must be invested to initiate a reaction is called its activation energy.
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All reactions have activation energy requirements, even extremely exergonic ones.
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In a chemical reaction, activation energy is the energy needed to put molecules into a transition state (i.e., to
turn them into what is called transition-state species).
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Transition-state species have higher free energy than either reactants or products. (See Figure 6.8.)
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Exergonic reactions often are initiated by the addition of heat (e.g., the lightning strike in a forest), which
increases the average kinetic energy of the molecules. (See Figure 6.9.) However, adding heat is not always an
appropriate way for biological systems to drive reactions.
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Enzymes, acting as biological catalysts, solve this problem. They substantially lower the required energy of
activation, but they do not initiate reactions that could not eventually take place on their own.
Enzymes bind specific reactant molecules
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Almost all enzymes are proteins, made of simple amino acids. Some enzymes also have important molecules
that participate in the catalysis, such as some types of vitamins and metal ions.
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Enzymes bind specific reactant molecules called substrates.
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Substrates bind to a particular site on the enzyme surface called the active site, where catalysis takes place.
(See Figure 6.10.)
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Enzymes are highly specific: They bind specific substrates and catalyze particular reactions under certain
conditions.
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An enzyme that catalyzes a certain reaction in one species might differ in amino acid composition from the
corresponding enzyme in another species, especially if the two species live at different temperatures and/or ionic
environments.
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The specificity of an enzyme comes from the three-dimensional shape of its active site.
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The amino acid sequence, temperature, and other conditions of the solution or the environment determine the
shape and structure of the active site.
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RNA polymerase catalyzes formation of RNA but not DNA.
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RNA nuclease hydrolyzes RNA polymers.
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Hexokinase accelerates phosphorylation of hexose. (All kinases add phosphates. All phosphatases remove
phosphates.)
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Binding a substrate to the active site produces an enzyme–substrate complex (ES).
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Hydrogen bonding, ionic attraction, or covalent bonding acting individually or together hold these complexes
together.
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The enzyme–substrate complex (ES) generates the product (P) and free enzyme (E): E + S  ES  E + P.
(See Figure 6.10.)
Enzymes lower the energy barrier but do not affect equilibrium
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Enzymes and other catalysts lower activation energy requirements and thus speed up the overall reaction, but
they do not change the difference in free energy (G) between the reactants and the products. Thus, they do not affect
the final equilibrium. (See Figure 6.11.)
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Enzymes can have a profound effect on rates toward equilibrium. Reactions that might take years to happen
can occur in a fraction of a second.
What are the chemical events at active sites of enzymes?
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At the active sites, enzymes and substrates interact by breaking old bonds and forming new ones. (See Figure
6.12.)
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Enzymes catalyze reactions using one or more of the following mechanisms: orienting substrates, adding
charges to substrates, or inducing strain in substrates.
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Enzymes orient substrates.
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While free in solution, substrates tumble and collide.
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The probability of collision at the angle necessary to change chemical interactions is low.
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When bound to enzymes, two substrates can be oriented such that a reaction is more likely to occur.
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Enzymes temporarily add chemical groups to substrates.
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The R groups (side chains) of an enzyme’s amino acids may participate directly in making substrates more
chemically reactive.
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In acid-base catalysis, acidic or basic side chains of amino acids form the active site and transfer H+ to or
from the substrate, destabilizing a covalent bond in a substrate.
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In covalent catalysis, a functional group side chain forms a temporary covalent bond with the substrate.
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In metal ion catalysis, metal ions gain or lose electrons without detaching from the protein, making them
important participants in oxygen-reduction (redox) reactions.
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Some enzymes induce strain in the substrate.
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For example, the carbohydrate substrate for the enzyme lysozyme enters the active site in a flat-ringed
“chair” shape. (See Figure 6.13.)
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The active site causes it to flatten out into a “sofa” shape.
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The stretching of the bonds decreases their stability, making them more reactive to water.
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(See Video 6.1.)
Molecular Structure Determines Enzyme Function
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Most enzymes (and ribozymes) are much larger than their substrate.
The active site of most enzymes is only a small region of the whole protein.
The active site is specific to the substrate
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The specificity of an enzyme for a particular substrate depends on a precise interlock.
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In 1894, Emil Fischer compared the fit to that of a lock and key, but his model was based only on indirect
evidence.
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In 1965, using X-ray crystallography, David Phillips observed a pocket in the enzyme lysozyme that neatly
fit its substrate. (See Figure 6.13.)
An enzyme changes shape when it binds a substrate
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The change in enzyme shape caused by substrate binding is called induced fit.
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Induced fit at least partly explains why enzymes are much larger than their substrates or reactants.
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Part of the larger size might be what allows induced fit. (See Figure 6.14.)
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Some regions of an enzyme tolerate little change without the enzyme’s losing its activity. This is the case
within the active site.
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Some enzymes can tolerate small changes in amino acid composition outside the active site and provide a
framework that enhances induced fit.
Some enzymes require other molecules in order to operate
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Cofactors are the inorganic ions in metal ion catalysis (e.g., copper, zinc, iron) that bind temporarily to
certain enzymes and are essential to their function.
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Coenzymes are carbon-containing molecules required for the action of one or more enzymes. (See Figure
6.15.)
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A coenzyme is like a substrate in that it is not permanently bound to the enzyme. It reacts with the enzyme,
binding to its active site. It changes chemically during the reaction, and then separates to participate in other reactions.
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ATP and ADP are coenzymes. In animals some coenzymes are produced from vitamins.
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Prosthetic groups are permanently bound to enzymes. They include the heme groups (iron-containing organic
molecules) that are attached to hemoglobin.
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Table 6.1 is a list of examples of cofactors, coenzymes, and prosthetic groups.
(See Video 6.2.)
Substrate concentration affects reaction rate
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The rate of an uncatalyzed reaction is directly proportional to the concentration of reactants. The higher the
substrate concentration, the more collisions and reactions per unit of time.
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This is true to a point with catalyzed reactions. At some point the enzyme will have all active sites occupied;
this is the point of saturation.
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Saturating an enzyme makes it possible to determine how many molecules are converted per unit time. (See
Figure 6.16.)
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The turnover number ranges from 1 molecule every 2 seconds for lysozyme, to 40 million per second for the
liver enzyme catalase.
Metabolism and the Regulation of Enzymes
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A major characteristic of life is homeostasis, the maintenance of stable internal conditions.
Regulation of enzyme activity contributes to homeostasis.
Metabolism is organized into pathways
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Metabolism is organized into sequences of enzyme-catalyzed chemical reactions called pathways.
A simple diagram of a biochemical pathway is the following:
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Each step (from A to B to C to D)
occurs appropriately because of enzymes. For
example, one enzyme converts A to B; a second enzyme converts B to C, and so on.
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Some metabolic pathways are anabolic and synthesize the building blocks of macromolecules.
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Some are catabolic and break down macromolecules and fuel molecules.
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In life, all reactions, taken together, are net exergonic, or energy releasing.
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All anabolic reactions are coupled to catabolic ones in a dynamic balance.
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The life we are most familiar with uses one important reaction to get the energy for the rest—the capture of a
photon of light into a chemical bond.
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Some other less familiar life-forms use the energy of methane or reduced inorganic substances as the starting
energy source for all other reactions.
Enzyme activity is subject to regulation by inhibitors
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Enzyme activity is inhibited by natural and artificial binders.
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Irreversible inhibition occurs when the inhibitor destroys the enzyme’s ability to interact with its normal
substrate(s).
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Diisopropylphosphorofluoridate, for example, reacts with the OH group of the serine residues found in active
sites, eliminating the activity of the enzyme. (See Figure 6.17.)
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Sarin, the nerve gas that was released in the Tokyo subway, is a related compound.
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Irreversible inhibitors are usually artificial, human-made compounds. Nature uses reversible inhibition to
regulate metabolism. (See Figure 6.18 and Animated Tutorial 6.1.)
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When an inhibitor binds reversibly to an enzyme’s active site, it competes with the substrate for the binding
site and is called a competitive inhibitor.
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When an inhibitor binds reversibly to a site away from the active site, it is called a noncompetitive inhibitor.
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Noncompetitive inhibitors act by changing the shape of the enzyme in such a way that the active site no
longer binds the substrate.
Allosteric enzymes control their activity by changing their shape
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The change in enzyme shape due to noncompetitive inhibitor binding is an example of allostery.
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Allosteric enzymes are controlled by allosteric regulators. (See Figure 6.19 and Animated Tutorial 6.2.)
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Allosteric regulators bind to an allosteric site, which is separate from the active site.
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This binding changes the structure and function of the enzyme.
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Some allosteric enzymes have multiple active sites.
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When one binding site is occupied, it changes the other(s) so that they bind additional substrate molecules
more readily.
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How the rate of an enzyme-catalyzed reaction changes with increasing substrate concentration depends on
whether the enzyme is allosterically regulated. (See Figure 6.20.)
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The advantage to the system is that the enzyme’s catalytic rate becomes concentration-sensitive and
concentration-responsive.
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Allosteric enzymes usually have more than one type of subunit.
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A catalytic subunit has an active site that binds the enzyme’s substrate.
A regulatory subunit has one or more allosteric sites that bind specific effector molecules.
An allosteric enzyme exists in an active or inactive form, similar to having an on/off switch.
In the active state, the active sites on the catalytic subunits can bind substrate.
In the inactive state, the allosteric sites on the regulatory subunits can accept the inhibitor.
The allosteric regulators that control these enzymes work in two ways:
Positive regulators stabilize the active form of the enzyme.
Negative regulators stabilize the inactive form of the enzyme.
(See Animated Tutorial 6.2.)
Allosteric effects regulate metabolism
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Metabolic pathways typically involve a starting material, intermediates, and a useful end-product.
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The first step in the pathway is called the start up or commitment step.
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Once this step occurs, other enzyme-catalyzed reactions follow until the product of the series builds up.
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One way to control the whole pathway is to have the end-product inhibit the first step in the pathway, the
commitment step.
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This is called end-product inhibition or feedback inhibition. (See Figure 6.21.)
Enzymes are affected by their environment
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Each enzyme is most active at a certain pH and temperature. (See Figures 6.22 and 6.23.)
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Some enzymes are tolerant of a wide range of pH and temperatures, while other enzymes are very sensitive.
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pH can influence the charges of carboxyl groups in neutral or basic solutions and amino groups in neutral or
acidic solutions.
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In general, an increase in temperature (to a point) increases the rate of an enzyme-catalyzed reaction. All
enzymes, as specific entities, show an optimal temperature for activity.
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Temperature can negatively influence shape by breaking hydrogen bonds and by interfering with ionic
interactions and hydrophobic interactions. If heat destroys the enzyme, the enzyme is denatured.
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Some organisms that can live at different temperatures generate different forms of an enzyme, called
isozymes.