contents Principles of Biology 23 ATP and Cellular Work ATP provides the energy that powers cells.

Magnetic resonance images of three different areas in the rat brain show blood flow and the biochemical measurements of ATP, pH, and glucose, which are all measures of energy use and production in brain tissue. The image is color-coded to show spatial differences in the concentration of these energy-related variables in brain tissue. © 1997 Nature Publishing Group Hoehn-Berlage, M., et al. Inhibition of nonselective cation channels reduces focal ischemic injury of rat brain. Journal of Cerebral Blood Flow and Metabolism 17, 534–542 (1997) doi: 10.1097/00004647-199705000-00007. Used with permission.

Topics Covered in this Module

Using Energy Resources For Work ATP-Driven Work

Major Objectives of this Module

Describe the role of ATP in energy-coupling reactions. Explain how ATP hydrolysis performs cellular work. Recognize chemical reactions that require ATP hydrolysis.

page 116 of 989 4 pages left in this module

contents Principles of Biology

23 ATP and Cellular Work

Energy is a fundamental necessity for all of life's processes. Without energy, flagella cannot move, DNA cannot be unwound or separated for replication or gene expression, cells cannot divide, plants cannot grow and animals cannot reproduce. Energy is vital, but where does it come from? Plants and photosynthetic microbes capture light energy and convert it into chemical energy for their own use. Organisms that cannot produce their own food, such as fungi and animals, feed upon this captured energy. However, the chemical energy produced by photosynthesizers needs to be converted into a usable form. Otherwise, it is like going to a grocery store with a bag of gold — until the gold is converted to the local currency, it is useless. For most organisms, that local currency is the molecule ATP.

Using Energy Resources For Work Many molecules can be used to store energy, but only one is called "the energy currency of the cell." What are the characteristics of a molecule that stores energy? How is this energy used by the cell to do work? Understanding the chemistry behind these questions will lead us to a more thorough understanding of how cells are able to conduct the business of living.

The molecule ATP functions as an energy carrier. Adenosine triphosphate (ATP) is composed of the nitrogenous base adenine, the sugar ribose, and three phosphate groups (Figure 1).

Figure 1: The structure of ATP. ATP is composed of the nitrogenous base adenine, a ribose sugar, and a chain of three phosphate groups. The unstable phosphoanhydride bonds permit transfer of phosphate groups to other molecules. © 2014 Nature Education All rights reserved.

Energy is stored in the phosphoanhydride bonds between the three phosphate groups in ATP. This energy is released by ATP hydrolysis, the chemical reaction in which ATP reacts with water to yield adenosine 2- diphosphate (ADP) and an inorganic phosphate ion, denoted HOPO3 or Pi (Figure 2). For every mole of ATP hydrolyzed, 7.3 kilocalories of energy are released:

ATP + H2O → ADP + Pi ΔG = -7.3 kcal/mol The standard Gibbs free energy (ΔG) is the portion of free energy that can be used by a system to perform work when temperature and pressure are uniform. It is a very handy number that can be used to predict whether a reaction will occur spontaneously within a cell. Exergonic reactions, such as ATP hydrolysis, release energy and have negative ΔG values. Conversely, endergonic reactions, which do not begin spontaneously and require an input of energy, have positive ΔG values.

Figure 2: The hydrolysis of ATP. A water molecule hydrolyzes ATP into ADP and inorganic phosphate (Pi 2- or HOPO3 ) by breaking the bond between the two terminal phosphate groups in ATP. The reaction also releases energy and a proton. Energy is released because the bond energy in the phosphoanhydride bond is greater than the energy in the bonds of the products. © 2013 Nature Education All rights reserved. A common misconception is that the phosphate bonds of ATP are high-energy bonds. However, there is nothing special about the bonds themselves. In reality, the energy release comes from the reaction between water and ATP, which results in the hydrolysis of the phosphate bonds. The reactants, water and ATP, have higher energy than the products, ADP and Pi, providing the energy commonly associated with ATP. But why are the products of ATP hydrolysis at lower energy than the reactants? ATP has three phosphate groups, all of which have negative charges. The close proximity of these charges destabilizes ATP because of the high degree of electrostatic repulsion. On the other hand, ADP has only two phosphate groups, which results in less electrostatic repulsion, lower energy, and therefore a more stable molecule. Furthermore, the other product of ATP hydrolysis also exhibits energetically favorable properties. Not only is Pi isolated from other negative charges, but it can be stabilized by interactions with water molecules in the cell (Figure 3). Taken together, Pi and ADP are at a much lower energy level than ATP. The hydrolysis of ATP releases this difference in energy. Figure 3: Ball and stick models of ATP, ADP and inorganic phosphate (Pi). ATP (a) is less chemically stable than ADP (b) because ADP has less electrostatic repulsion between negatively charged phosphate groups. Inorganic phosphate (c) has four oxygen atoms with partial negative charges that can be stabilized by water molecules in biological systems. These qualities make ADP and Pi much more stable than ATP. Red, blue, gray, white and orange spheres represent oxygen, nitrogen, carbon, hydrogen and phosphorus atoms, respectively. © 2013 Nature Education All rights reserved.

ATP powers energetically unfavorable reactions in the cell. How does the hydrolysis of ATP lead to the cell carrying out useful reactions? The release of energy only explains half the story. On its own, the hydrolysis of ATP would simply result in organisms overheating because the dissipation of energy would excite nearby molecules, resulting in heat (thermal energy). For energy to be useful in a cell, it needs to be linked to other processes. Energy coupling is the transfer of energy from one chemical reaction to another. By coupling the release of energy that occurs during ATP hydrolysis with changes in protein conformation, the breaking and forming of chemical bonds, and other reactions that require energy, the cell can perform nearly all of the tasks it needs to function.

How is ATP used during energy-coupling reactions? As an example, consider the phosphorylation of the sugar glucose into glucose-6-phosphate (Figure 4). This is the first step in glycolysis, one of the most fundamental processes for generating ATP in all organisms. The direct addition of an inorganic phosphate group to glucose will not occur spontaneously because it involves a positive free energy change (ΔG > 0) — it is endergonic. To overcome this problem, cells couple glucose phosphorylation to ATP hydrolysis through a two-step process catalyzed by the enzyme hexokinase. First, ATP is hydrolyzed into ADP and an inorganic phosphate group. The hydrolysis of ATP is exergonic, involving a negative free energy change (ΔG < 0). The inorganic phosphate group released from the first reaction is then transferred to glucose, forming glucose-6-phosphate. This initial investment of energy is needed to completely break down glucose in later steps of glycolysis, which release more net energy.

Figure 4: The phosphorylation of glucose is coupled with ATP hydrolysis. In this reaction, the addition of an inorganic phosphate group (Pi) to glucose to form glucose-6-phosphate — the first step of glycolysis — is energetically unfavorable (∆G = +3.3 kcal/mol). The hydrolysis of ATP into ADP and Pi is energetically favorable (-7.3 kcal/mol). Coupling the two reactions allows the energy released during ATP hydrolysis to be used to drive the endergonic reaction. © 2014 Nature Education All rights reserved. Figure Detail The ΔG of the direct phosphorylation of glucose is +3.3 kcal/mol, and the ΔG of ATP hydrolysis is -7.3 kcal/mol. By coupling these reactions, the ∆G values of each reaction are summed, giving an overall ∆G of -4.0 kcal/mol (+3.3 kcal/mol + (-7.3 kcal/mol) = -4.0 kcal/mol). Because the free energy change of the overall reaction is negative, the overall reaction is energetically favorable and will occur spontaneously. In this way, ATP hydrolysis can be used to provide the free energy needed to drive reactions that would otherwise be energetically unfavorable (Figure 5).

Figure 5: Spontaneous reactions. Chemical reactions will occur spontaneously if the free energy of the reaction is negative. © 2013 Nature Education All rights reserved. Transcript

The cycling of ATP. The exergonic hydrolysis of ATP is coupled to many, but not all, endergonic reactions in the cell. It fuels the movement of muscles, the movement of molecules into and out of cells, chemical reactions, and more. Humans use approximately 50 kg of ATP each day. However, not even Olympic sprinters can store 50 kg of ATP in their muscles; at most, they store approximately one minute's worth of ATP. Obviously, ATP must be regenerated continuously. The addition of an inorganic phosphate to ADP regenerates ATP. This reaction, the phosphorylation of ADP, is endergonic and requires +7.3 kcal/mol of energy. (Note that the opposite reaction, the hydrolysis of ATP, releases the same amount of energy.)

ADP + Pi → ATP + H2O ΔG = +7.3 kcal/mol For most life forms, the energy required to regenerate ATP comes from respiration of the chemical energy initially stored during photosynthesis; this stored energy is used by either the organism that did the photosynthesis itself or an organism that consumed the products of that photosynthesis. Some of the energy released by respiration can be used to synthesize ATP. In turn, the hydrolysis of ATP is coupled with energy-costing processes (Figure 6). The regeneration of ATP is very quick and efficient. For example, human muscle cells use and regenerate approximately 10 million molecules of ATP per second per cell! Figure 6: The ATP cycle.Energy from catabolism or the capture of light energy is used to drive ATP synthesis from ADP and Pi. In turn, ATP is hydrolyzed back into ADP and Pi to provide energy for cellular processes. © 2013 Nature Education All rights reserved.

IN THIS MODULE

Using Energy Resources For Work ATP-Driven Work Summary Test Your Knowledge

PRIMARY LITERATURE

Mitochondria change shape to help the cell survive During autophagy mitochondria elongate, are spared from degradation and sustain cell viability. View | Download

How can nematodes help reduce obesity in humans? A whole-organism screen identifies new regulators of fat storage. View | Download

SCIENCE ON THE WEB

Cells and Energy Learn more about ATP synthase from John Walker's lab webpage.

ATP Wins the Prize Read about the Nobel Prize winners in Chemistry for their contribution to ATP studies

Energy in Action CSU professor Sandra Jewett has compiled a list of useful links about ATP synthase.

page 117 of 989 3 pages left in this module

contents Principles of Biology

23 ATP and Cellular Work ATP-Driven Work Cells need energy to grow, reproduce and maintain homeostasis, but for the energy provided by ATP to be useful, it must be coupled with work. In a cell, this work takes three main forms — chemical work, mechanical work, and transport work. Glutamine synthesis is an example of how ATP hydrolysis is used to perform chemical work by driving unfavorable chemical processes that would not occur spontaneously. Another example is in photosynthesis itself, where carbon atoms in carbon dioxide are bonded together to form sugars. ATP must be hydrolyzed to provide the energy to form these new carbon-carbon bonds. Any anabolic activity in a cell will likely require chemical work and therefore ATP consumption.

How does ATP help a cell perform mechanical work? In addition to chemical work, ATP can also power mechanical work. Helicases are a class of proteins that separate double-stranded nucleic acids, most notably double-stranded DNA, into single strands. This process requires a large amount of energy, which is supplied by ATP hydrolysis. To separate double-stranded DNA, DNA helicase moves along a strand of DNA and individually breaks the hydrogen bonds between complementary bases, one at a time, leaving two single strands of DNA. Each breakage event is fueled by ATP hydrolysis (Figure 7).

Figure 7: Helicase unwinds DNA. DNA helicase is a protein that changes conformation, in essence "walking" along the DNA and separating the two strands. Repeated ATP hydrolysis powers the changes in shape that allow DNA helicase to move. © 2013 Nature Education All rights reserved.

ATP also fuels mechanical work in other types of cells. An important example in humans is muscle contraction, which is caused by the sliding of actin and myosin filaments past each other in muscle cells. In this case, ATP hydrolysis allows the cyclical interaction between actin and myosin to continue during a contraction. Without ATP, muscles remain trapped in a contracted state. This explains the phenomenon of rigor mortis; after death, the absence of ATP prevents actin and myosin filaments from separating from each other.

How does ATP drive membrane transport? ATP is also used to power the transport of substances across cell membranes and against their concentration gradients. In some cases, ATP is used to transport one type of substance in one direction (see Figure 8). In other cases, two substances are moved in opposite directions using energy provided by ATP. For example, neurons rely on high concentrations of sodium (Na+) ions outside the cell and potassium (K+) ions inside the cell. The accumulation of these ions against their concentration gradients is mediated by the sodium-potassium pump, or Na+/K+ ATPase. As implied in its name, the transporter hydrolyzes ATP to pump three Na+ ions out of the cell and two K+ ions into the cell. First, the Na+ ions bind to the pump, followed by ATP. The ATP is hydrolyzed, providing the energy to change the conformation of the pump from inward-facing to outward-facing, leaving inorganic phosphate bound to the pump. The now outward-facing pump releases the Na+ ions out of the cell and allows the K+ ions to bind the protein pump. A final conformational change, from outward- to inward-facing, is caused by the release of the phosphate group, which releases the K+ ions into the cell.

Figure 8: Active transport. Some proteins require energy to transport ions across the cell membrane.

© 2014 Nature Education All rights reserved. Transcript

The activity of the sodium-potassium pump creates a high concentration of Na+ outside the cell. Upon stimulation, Na+ rapidly flows back into the cell, changing the ion concentrations across the cell membrane and producing an electrical signal. The sodium-potassium pump is so central to neural function that the Danish chemist Jens Christian Skou received the Nobel Prize in Chemistry in 1997 for his work in uncovering this mechanism. Test Yourself

DNA ligase is an enzyme that repairs gaps in the DNA phosphate backbone by forming new phosphodiester bonds. Would this process likely require ATP hydrolysis? If so, what form of cellular work is being performed? Explain.

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Is ATP the only "energy molecule"? ATP is not the only energy carrier within cells. For example, creatine phosphate can release 9.6 kcal/mol, and phosphoenolpyruvate can release a whopping 14.8 kcal/mol. If phosphoenolpyruvate can release almost twice as much energy as ATP, then why hasn't phosphoenolpyruvate replaced ATP as the cell's main energy currency through evolutionary time? Efficiency is the key. Energy carriers are analogous to money in a system that does not give change. When a cell "pays" more for a reaction than the reaction requires, any leftover energy is released as heat. Suppose a particular endergonic reaction has a ΔG of +4.8 kcal/mol. If the cell pays for this reaction by hydrolyzing ATP, an exergonic reaction with a negative ΔG of -7.3 kcal/mol, then the free energy difference of -2.5 kcal/mol will be released as heat. If the cell pays for this reaction by hydrolyzing phosphoenolpyruvate, an exergonic reaction that releases much more energy (ΔG = -14.8 kcal/mol), then -10 kcal/mol of energy — nearly four times as much than if ATP were used — is released as heat instead. In other words, the higher-value currency will not result in more work being done; it just means that more energy will be released as heat, rather than being used for basic functions that sustain life. The beauty behind ATP is its utility, ease of use, and rapidity of regeneration. Most reactions require less energy than the hydrolysis of ATP releases, so ATP is a very efficient energy currency.

CAREERS ATP Researchers May Be Biologists, Chemists, or Physicists ATP researchers often straddle different disciplines of science, depending on which aspect of ATP they study. Those researching the molecular structure, reactivity, and energetics of ATP may be working largely in chemistry and physics labs. Those studying how cells use ATP likely work in a biology lab, perhaps with cell cultures and a range of microscopy equipment. Regardless of the academic discipline in which the research is conducted, the study of ATP has been a treasure trove for scientists. In 1997, John Ernest Walker and Paul D. Boyer were awarded the Nobel Prize in Chemistry for uncovering how ATP is regenerated. Walker determined the structure of ATP synthase, which makes ATP, down to an atomic-level resolution. He used bovine heart mitochondria and eubacteria for his experiments, which highlighted how well conserved and essential this protein is. Boyer cleverly showed how the energy released by catabolism is converted to useful energy to generate ATP from ADP and Pi. In his key experiments, Boyer tracked radioactively labeled oxygen as it was turned into radioactive water by ATP synthase. Despite ATP being one of the most studied molecules in biology, scientists still have many questions to answer about how the molecule functions and how it is regenerated. A number of labs are actively examining topics such as the exact mechanisms of ATP synthase as well as the importance of similarities and differences in ATP synthesis and utilization across species.

CAREERS

IN THIS MODULE Using Energy Resources For Work ATP-Driven Work Summary Test Your Knowledge

PRIMARY LITERATURE

Mitochondria change shape to help the cell survive During autophagy mitochondria elongate, are spared from degradation and sustain cell viability. View | Download

How can nematodes help reduce obesity in humans? A whole-organism screen identifies new regulators of fat storage. View | Download

SCIENCE ON THE WEB

Cells and Energy Learn more about ATP synthase from John Walker's lab webpage.

ATP Wins the Prize Read about the Nobel Prize winners in Chemistry for their contribution to ATP studies

Energy in Action CSU professor Sandra Jewett has compiled a list of useful links about ATP synthase.

page 118 of 989 2 pages left in this module

contents Principles of Biology

23 ATP and Cellular Work Summary OBJECTIVE Describe the role of ATP in energy-coupling reactions. The ATP molecule is composed of a nitrogenous base called adenine, a ribose sugar and a chain of three phosphate groups. Having three phosphate groups makes ATP relatively unstable when compared to ADP and inorganic 2- phosphate (Pi or HOPO3 ), which are the products of its hydrolysis. This difference in stability between products and reactants causes ATP hydrolysis to release a large amount of energy, which can be coupled to energetically unfavorable reactions in the cell.

OBJECTIVE Recognize chemical reactions that require ATP hydrolysis. ATP can drive other reactions because its hydrolysis is exergonic and releases energy. Therefore, the hydrolysis of ATP is spontaneous, having a negative free energy change (-ΔG). Spontaneous reactions can drive non-spontaneous reactions if they are coupled together.

OBJECTIVE Explain how ATP hydrolysis performs cellular work. There are three main types of cellular work: chemical, mechanical, and transport. For example, ATP can be used to provide the mechanical work that powers DNA helicase to separate double-stranded DNA into single- stranded DNA during DNA replication. One of the most vital roles of ATP is the coupling of ATP hydrolysis to active transport of ions across cell membranes.

Key Terms adenosine triphosphate (ATP) The primary energy currency of the cell; composed of the nitrogenous base adenine, the 5-carbon sugar ribose and three phosphate groups.

ATP hydrolysis The exergonic chemical reaction in which the combination of ATP with water results in the release of an inorganic phosphate ion and ADP.

endergonic reaction A chemical reaction with a net requirement for input of energy.

energy coupling Transfer of energy from one chemical reaction to another process. exergonic reaction A chemical reaction with a net requirement for input of energy. phosphoanhydride bond Covalent bond that links two phosphate groups to one another. Two are found in ATP. Hydrolysis of one of these bonds releases 7.3 kcal/mol of energy.

IN THIS MODULE

Using Energy Resources For Work ATP-Driven Work Summary Test Your Knowledge

PRIMARY LITERATURE

Mitochondria change shape to help the cell survive During autophagy mitochondria elongate, are spared from degradation and sustain cell viability. View | Download

How can nematodes help reduce obesity in humans? A whole-organism screen identifies new regulators of fat storage. View | Download

SCIENCE ON THE WEB

Cells and Energy Learn more about ATP synthase from John Walker's lab webpage.

ATP Wins the Prize Read about the Nobel Prize winners in Chemistry for their contribution to ATP studies

Energy in Action CSU professor Sandra Jewett has compiled a list of useful links about ATP synthase.

page 119 of 989 1 pages left in this module

contents Principles of Biology

23 ATP and Cellular Work

Test Your Knowledge

1. Identify the structural features found in ATP.

a nitrogenous base, a deoxyribose sugar, and a phosphate group a nitrogenous base, a ribose sugar, and a phosphate group a nitrogenous base, a ribose sugar, and three phosphate groups a nucleoside, a ribose sugar, and three phosphate groups a nitrogenous base, a ribose sugar, and two phosphate groups

2. Why does ATP hydrolysis release a lot of free energy?

ATP is unstable compared to its products. ATP has a nitrogenous base. ATP has special high-energy phosphate bonds. ATP has a ribose sugar. The products of ATP are less stable than ATP itself.

3. ATP is not the only molecule that can drive reactions. When ATP is depleted during exercise, muscle cells use phosphocreatine to drive the regeneration of ATP.

Consider the following half reactions:

1. ATP + H2O → ADP + Pi

ΔG = –7.3 kcal/mol

2. Phosphocreatine + H2O → creatine + Pi

ΔG = –10.3 kcal/mol

From these two reactions, calculate the Gibbs free energy of the following coupled reaction, catalyzed by creatine kinase:

Phosphocreatine + ADP → ATP + creatine

ΔG = ?

Which is the correct net Gibbs free energy of the reaction?

ΔG = –3 kcal/mol ΔG = –17.6 kcal/mol ΔG = 0 kcal/mol ΔG = +3 kcal/mol ΔG = +17.6 kcal/mol

4. The sodium-potassium pump is an active transport pump that uses energy to pump potassium into cells and sodium out of cells. Why is ATP energy required?

to change the shape of the ions to transfer glucose in the same direction as Na+ ions to pump the Na+ and K+ ions along their diffusion gradient to pump the Na+ and K+ ions against their diffusion gradient for the membrane to change shape

5. Complete the following sentence: The hydrolysis of one mole of ATP can be used to drive reactions that have a ΔG that is... less than 7.3 kcal/mol. more than 7.3 kcal/mol. less than –7.3 kcal/mol. exactly 7.3 kcal/mol. exactly –7.3 kcal/mol.

Submit

IN THIS MODULE

Using Energy Resources For Work ATP-Driven Work Summary Test Your Knowledge

PRIMARY LITERATURE

Mitochondria change shape to help the cell survive During autophagy mitochondria elongate, are spared from degradation and sustain cell viability. View | Download

How can nematodes help reduce obesity in humans? A whole-organism screen identifies new regulators of fat storage. View | Download

SCIENCE ON THE WEB

Cells and Energy Learn more about ATP synthase from John Walker's lab webpage.

ATP Wins the Prize Read about the Nobel Prize winners in Chemistry for their contribution to ATP studies

Energy in Action CSU professor Sandra Jewett has compiled a list of useful links about ATP synthase.

page 120 of 989