7.2: Catabolism of Carbohydrates

Learning Objectives

• Describe why glycolysis is not oxygen dependent
• Define and describe the net yield of three-carbon molecules, ATP, and NADH from glycolysis
• Explain how three-carbon pyruvate molecules are converted into two-carbon acetyl groups that can be funneled into the Krebs cycle.
• Define and describe the net yield of CO₂, GTP/ATP, FADH₂, and NADH from the Krebs cycle
• Explain how intermediate carbon molecules of the Krebs cycle can be used in a cell
• Compare and contrast the electron transport system location and function in a prokaryotic cell and a eukaryotic cell
• Compare and contrast the differences between substrate-level and oxidative phosphorylation
• Explain the relationship between chemiosmosis and proton motive force
• Describe the function and location of ATP synthase in a prokaryotic versus eukaryotic cell
• Compare and contrast aerobic and anaerobic respiration

Extensive enzyme pathways exist for breaking down carbohydrates to capture energy in ATP bonds. In addition, many catabolic pathways produce intermediate molecules that are also used as building blocks for anabolic reactions. Understanding these processes is important for several reasons. First, because the main metabolic processes involved are common to a wide range of chemoheterotrophic organisms, we can learn a great deal about human metabolism by studying metabolism in more easily manipulated bacteria like E. coli. Second, because animal and human pathogens are also chemoheterotrophs, learning about the details of metabolism in these bacteria, including possible differences between bacterial and human pathways, is useful for the diagnosis of pathogens as well as for the discovery of antimicrobial therapies targeting specific pathogens. Last, learning specifically about the pathways involved in chemoheterotrophic metabolism also serves as a basis for comparing other more unusual metabolic strategies used by microbes. Although the chemical source of electrons initiating electron transfer is different between chemoheterorophs and chemoautotrophs, many similar processes are used in both types of organisms.

The typical example used to introduce concepts of metabolism to students is carbohydrate catabolism. For chemoheterotrophs, our examples of metabolism start with the catabolism of polysaccharides such as glycogen, starch, or cellulose. Enzymes such as amylase, which breaks down glycogen or starch, and cellulases, which break down cellulose, can cause the hydrolysis of glycosidic bonds between the glucose monomers in these polymers, releasing glucose for further catabolism.

Glycolysis

For bacteria, eukaryotes, and most archaea, glycolysis is the most common pathway for the catabolism of glucose; it produces energy, reduced electron carriers, and precursor molecules for cellular metabolism. Every living organism carries out some form of glycolysis, suggesting this mechanism is an ancient universal metabolic process. The process itself does not use oxygen; however, glycolysis can be coupled with additional metabolic processes that are either aerobic or anaerobic. Glycolysis takes place in the cytoplasm of prokaryotic and eukaryotic cells. It begins with a single six-carbon glucose molecule and ends with two molecules of a three-carbon sugar called pyruvate. Pyruvate may be broken down further after glycolysis to harness more energy through aerobic or anaerobic respiration, but many organisms, including many microbes, may be unable to respire; for these organisms, glycolysis may be their only source of generating ATP.

The type of glycolysis found in animals and that is most common in microbes is the Embden-Meyerhof-Parnas (EMP) pathway, named after Gustav Embden (1874–1933), Otto Meyerhof (1884–1951), and Jakub Parnas (1884–1949). Glycolysis using the EMP pathway consists of two distinct phases (Figure \(\PageIndex{1}\)). The first part of the pathway, called the energy investment phase, uses energy from two ATP molecules to modify a glucose molecule so that the six-carbon sugar molecule can be split evenly into two phosphorylated three-carbon molecules called glyceraldehyde 3-phosphate (G3P). The second part of the pathway, called the energy payoff phase, extracts energy by oxidizing G3P to pyruvate, producing four ATP molecules and reducing two molecules of NAD⁺ to two molecules of NADH, using electrons that originated from glucose.

The ATP molecules produced during the energy payoff phase of glycolysis are formed by substrate-level phosphorylation (Figure \(\PageIndex{1}\)), one of two mechanisms for producing ATP. In substrate-level phosphorylation, a phosphate group is removed from an organic molecule and is directly transferred to an available ADP molecule, producing ATP. During glycolysis, high-energy phosphate groups from the intermediate molecules are added to ADP to make ATP.

Overall, in this process of glycolysis, the net gain from the breakdown of a single glucose molecule is:

• two ATP molecules
• two NADH molecule, and
• two pyruvate molecules.

Figure \(\PageIndex{1}\): The energy investment phase of the Embden-Meyerhof-Parnas glycolysis pathway uses two ATP molecules to phosphorylate glucose, forming two glyceraldehyde 3-phosphate (G3P) molecules. The energy payoff phase harnesses the energy in the G3P molecules, producing four ATP molecules, two NADH molecules, and two pyruvates.

Figure \(\PageIndex{2}\): The ATP made during glycolysis is a result of substrate-level phosphorylation. One of the two enzymatic reactions in the energy payoff phase of Embden Meyerhof-Parnas glycolysis that produce ATP in this way is shown here.

Other Glycolytic Pathways

When we refer to glycolysis, unless otherwise indicated, we are referring to the EMP pathway used by animals and many bacteria. However, some prokaryotes use alternative glycolytic pathways. One important alternative is the Entner-Doudoroff (ED) pathway, named after its discoverers Nathan Entner and Michael Doudoroff (1911–1975). Although some bacteria, including the opportunistic gram-negative pathogen Pseudomonas aeruginosa, contain only the ED pathway for glycolysis, other bacteria, like E. coli, have the ability to use either the ED pathway or the EMP pathway.

A third type of glycolytic pathway that occurs in all cells, which is quite different from the previous two pathways, is the pentose phosphate pathway (PPP) also called the phosphogluconate pathway or the hexose monophosphate shunt. Evidence suggests that the PPP may be the most ancient universal glycolytic pathway. The intermediates from the PPP are used for the biosynthesis of nucleotides and amino acids. Therefore, this glycolytic pathway may be favored when the cell has need for nucleic acid and/or protein synthesis, respectively.

Exercise \(\PageIndex{1}\)

When might an organism use the ED pathway or the PPP for glycolysis?

Transition Reaction, Coenzyme A, and the Krebs Cycle

Glycolysis produces pyruvate, which can be further oxidized to capture more energy. For pyruvate to enter the next oxidative pathway, it must first be decarboxylated by the enzyme complex pyruvate dehydrogenase to a two-carbon acetyl group in the transition reaction, also called the bridge reaction (Figure \(\PageIndex{3}\)). In the transition reaction, electrons are also transferred to NAD⁺ to form NADH. To proceed to the next phase of this metabolic process, the comparatively tiny two-carbon acetyl must be attached to a very large carrier compound called coenzyme A (CoA). The transition reaction occurs in the mitochondrial matrix of eukaryotes; in prokaryotes, it occurs in the cytoplasm because prokaryotes lack membrane-enclosed organelles.

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Figure \(\PageIndex{3}\): Upon entering the mitochondrial matrix, a multienzyme complex converts pyruvate into acetyl CoA. In the process, carbon dioxide is released, and one molecule of NADH is formed.

The Krebs cycle transfers remaining electrons from the acetyl group produced during the transition reaction to electron carrier molecules, thus reducing them. The Krebs cycle also occurs in the cytoplasm of prokaryotes along with glycolysis and the transition reaction, but it takes place in the mitochondrial matrix of eukaryotic cells where the transition reaction also occurs. The Krebs cycle is named after its discoverer, British scientist Hans Adolf Krebs (1900–1981) and is also called the citric acid cycle, or the tricarboxylic acid cycle (TCA) because citric acid has three carboxyl groups in its structure. Unlike glycolysis, the Krebs cycle is a closed loop: The last part of the pathway regenerates the compound used in the first step (Figure \(\PageIndex{4}\)). The eight steps of the cycle are a series of chemical reactions that capture the two-carbon acetyl group (the CoA carrier does not enter the Krebs cycle) from the transition reaction, which is added to a four-carbon intermediate in the Krebs cycle, producing the six-carbon intermediate citric acid (giving the alternate name for this cycle). As one turn of the cycle returns to the starting point of the four-carbon intermediate, the cycle produces two CO₂ molecules, one ATP molecule (or an equivalent, such as guanosine triphosphate [GTP]) produced by substrate-level phosphorylation, and three molecules of NADH and one of FADH₂.

Figure \(\PageIndex{4}\): (a) Coenzyme A is shown here without an attached acetyl group. (b) Coenzyme A is shown here with an attached acetyl group.

Although many organisms use the Krebs cycle as described as part of glucose metabolism, several of the intermediate compounds in the Krebs cycle can be used in synthesizing a wide variety of important cellular molecules, including amino acids, chlorophylls, fatty acids, and nucleotides; therefore, the cycle is both anabolic and catabolic (Figure \(\PageIndex{5}\)).

Figure \(\PageIndex{5}\): The Krebs cycle, also known as the citric acid cycle, is summarized here. Note incoming two-carbon acetyl results in the main outputs per turn of two CO₂, three NADH, one FADH₂, and one ATP (or GTP) molecules made by substrate-level phosphorylation. Two turns of the Krebs cycle are required to process all of the carbon from one glucose molecule.

Figure \(\PageIndex{6}\): Many organisms use intermediates from the Krebs cycle, such as amino acids, fatty acids, and nucleotides, as building blocks for biosynthesis.

The Last Step

We have just discussed two pathways in glucose catabolism—glycolysis and the Krebs cycle—that generate ATP by substrate-level phosphorylation. Most ATP, however, is generated during a separate process called oxidative phosphorylation, which occurs during cellular respiration. Cellular respiration begins when electrons are transferred from NADH and FADH₂—made in glycolysis, the transition reaction, and the Krebs cycle—through a series of chemical reactions to a final inorganic electron acceptor (either oxygen in aerobic respiration or non-oxygen inorganic molecules in anaerobic respiration). These electron transfers take place on the inner part of the cell membrane of prokaryotic cells or in specialized protein complexes in the inner membrane of the mitochondria of eukaryotic cells. The energy of the electrons is harvested to generate an electrochemical gradient across the membrane, which is used to make ATP by oxidative phosphorylation.

Electron Transport System

The electron transport chain (ETC) or electron transport system (ETS) is the last component involved in the process of cellular respiration; it comprises a series of membrane-associated protein complexes and associated mobile accessory electron carriers. Electron transport is a series of chemical reactions that resembles a bucket brigade in that electrons from NADH and FADH₂ are passed rapidly from one ETS electron carrier to the next. These carriers can pass electrons along in the ETS because of their redox potential. For a protein or chemical to accept electrons, it must have a more positive redox potential than the electron donor. Therefore, electrons move from electron carriers with more negative redox potential to those with more positive redox potential. The four major classes of electron carriers involved in both eukaryotic and prokaryotic electron transport systems are the cytochromes, flavoproteins, iron-sulfur proteins, and the quinones.

In aerobic respiration, the final electron acceptor (i.e., the one having the most positive redox potential) at the end of the ETS is an oxygen molecule (O₂) that becomes reduced to water (H₂O) by the final ETS carrier. This electron carrier, cytochrome oxidase, differs between bacterial types and can be used to differentiate closely related bacteria for diagnoses. For example, the gram-negative opportunist Pseudomonas aeruginosa and the gram-negative cholera-causing Vibrio cholerae use cytochrome c oxidase, which can be detected by the oxidase test, whereas other gram-negative Enterobacteriaceae, like E. coli, are negative for this test because they produce different cytochrome oxidase types.

There are many circumstances under which aerobic respiration is not possible, including any one or more of the following:

• The cell lacks genes encoding an appropriate cytochrome oxidase for transferring electrons to oxygen at the end of the electron transport system.
• The cell lacks genes encoding enzymes to minimize the severely damaging effects of dangerous oxygen radicals produced during aerobic respiration, such as hydrogen peroxide (H₂O₂) or superoxide (O2–).
• The cell lacks a sufficient amount of oxygen to carry out aerobic respiration.

One possible alternative to aerobic respiration is anaerobic respiration, using an inorganic molecule other than oxygen as a final electron acceptor. There are many types of anaerobic respiration found in bacteria and archaea. Denitrifiers are important soil bacteria that use nitrate (NO3–) and nitrite (NO2–) as final electron acceptors, producing nitrogen gas (N₂). Many aerobically respiring bacteria, including E. coli, switch to using nitrate as a final electron acceptor and producing nitrite when oxygen levels have been depleted.

Microbes using anaerobic respiration commonly have an intact Krebs cycle, so these organisms can access the energy of the NADH and FADH₂ molecules formed. However, anaerobic respirers use altered ETS carriers encoded by their genomes, including distinct complexes for electron transfer to their final electron acceptors. Smaller electrochemical gradients are generated from these electron transfer systems, so less ATP is formed through anaerobic respiration.

Exercise \(\PageIndex{2}\)

Do both aerobic respiration and anaerobic respiration use an electron transport chain?

Chemiosmosis, Proton Motive Force, and Oxidative Phosphorylation

In each transfer of an electron through the ETS, the electron loses energy, but with some transfers, the energy is stored as potential energy by using it to pump hydrogen ions (H⁺) across a membrane. In prokaryotic cells, H⁺ is pumped to the outside of the cytoplasmic membrane (called the periplasmic space in gram-negative and gram-positive bacteria), and in eukaryotic cells, they are pumped from the mitochondrial matrix across the inner mitochondrial membrane into the intermembrane space. There is an uneven distribution of H⁺ across the membrane that establishes an electrochemical gradient because H⁺ ions are positively charged (electrical) and there is a higher concentration (chemical) on one side of the membrane. This electrochemical gradient formed by the accumulation of H⁺ (also known as a proton) on one side of the membrane compared with the other is referred to as the proton motive force (PMF). Because the ions involved are H⁺, a pH gradient is also established, with the side of the membrane having the higher concentration of H⁺ being more acidic. Beyond the use of the PMF to make ATP, as discussed in this chapter, the PMF can also be used to drive other energetically unfavorable processes, including nutrient transport and flagella rotation for motility.

The potential energy of this electrochemical gradient generated by the ETS causes the H⁺ to diffuse across a membrane (the plasma membrane in prokaryotic cells and the inner membrane in mitochondria in eukaryotic cells). This flow of hydrogen ions across the membrane, called chemiosmosis, must occur through a channel in the membrane via a membrane-bound enzyme complex called ATP synthase (Figure \(\PageIndex{7}\)). The tendency for movement in this way is much like water accumulated on one side of a dam, moving through the dam when opened. ATP synthase (like a combination of the intake and generator of a hydroelectric dam) is a complex protein that acts as a tiny generator, turning by the force of the H⁺ diffusing through the enzyme, down their electrochemical gradient from where there are many mutually repelling H⁺ to where there are fewer H⁺. In prokaryotic cells, H⁺ flows from the outside of the cytoplasmic membrane into the cytoplasm, whereas in eukaryotic mitochondria, H⁺ flows from the intermembrane space to the mitochondrial matrix. The turning of the parts of this molecular machine regenerates ATP from ADP and inorganic phosphate (P_i) by oxidative phosphorylation, a second mechanism for making ATP that harvests the potential energy stored within an electrochemical gradient.

Figure \(\PageIndex{7}\): The electron transport chain is a series of electron carriers and ion pumps that are used to pump H⁺ ions across a membrane. H⁺ then flow back through the membrane by way of ATP synthase, which catalyzes the formation of ATP. The location of the electron transport chain is the inner mitochondrial matrix in eukaryotic cells and cytoplasmic membrane in prokaryotic cells.

The number of ATP molecules generated from the catabolism of glucose varies. For example, the number of hydrogen ions that the electron transport system complexes can pump through the membrane varies between different species of organisms. In aerobic respiration in mitochondria, the passage of electrons from one molecule of NADH generates enough proton motive force to make three ATP molecules by oxidative phosphorylation, whereas the passage of electrons from one molecule of FADH₂ generates enough proton motive force to make only two ATP molecules. Thus, the 10 NADH molecules made per glucose during glycolysis, the transition reaction, and the Krebs cycle carry enough energy to make 30 ATP molecules, whereas the two FADH₂ molecules made per glucose during these processes provide enough energy to make four ATP molecules. Overall, the theoretical maximum yield of ATP made during the complete aerobic respiration of glucose is 38 molecules, with four being made by substrate-level phosphorylation and 34 being made by oxidative phosphorylation (Figure \(\PageIndex{8}\)). In reality, the total ATP yield is usually less, ranging from one to 34 ATP molecules, depending on whether the cell is using aerobic respiration or anaerobic respiration; in eukaryotic cells, some energy is expended to transport intermediates from the cytoplasm into the mitochondria, affecting ATP yield. Figure \(\PageIndex{8}\) summarizes the theoretical maximum yields of ATP from various processes during the complete aerobic respiration of one glucose molecule.

Figure \(\PageIndex{8}\): Theoretical maximum yields of ATP from various processes during the complete aerobic respiration of one glucose molecule.

Exercise \(\PageIndex{3}\)

What are the functions of the proton motive force?

Clinical Focus: part 2

Many of Hannah’s symptoms are consistent with several different infections, including influenza and pneumonia. However, her sluggish reflexes along with her light sensitivity and stiff neck suggest some possible involvement of the central nervous system, perhaps indicating meningitis. Meningitis is an infection of the cerebrospinal fluid (CSF) around the brain and spinal cord that causes inflammation of the meninges, the protective layers covering the brain. Meningitis can be caused by viruses, bacteria, or fungi. Although all forms of meningitis are serious, bacterial meningitis is particularly serious. Bacterial meningitis may be caused by several different bacteria, but the bacterium Neisseria meningitidis, a gram-negative, bean-shaped diplococcus, is a common cause and leads to death within 1 to 2 days in 5% to 10% of patients.

Given the potential seriousness of Hannah’s conditions, her physician advised her parents to take her to the hospital in the Gambian capital of Banjul and there have her tested and treated for possible meningitis. After a 3-hour drive to the hospital, Hannah was immediately admitted. Physicians took a blood sample and performed a lumbar puncture to test her CSF. They also immediately started her on a course of the antibiotic ceftriaxone, the drug of choice for treatment of meningitis caused by N. meningitidis, without waiting for laboratory test results.

Exercise \(\PageIndex{4}\)

1. How might biochemical testing be used to confirm the identity of N. meningitidis?
2. Why did Hannah’s doctors decide to administer antibiotics without waiting for the test results?

Key Concepts and Summary

• Glycolysis is the first step in the breakdown of glucose, resulting in the formation of ATP, which is produced by substrate-level phosphorylation; NADH; and two pyruvate molecules. Glycolysis does not use oxygen and is not oxygen dependent.
• After glycolysis, a three-carbon pyruvate is decarboxylated to form a two-carbon acetyl group, coupled with the formation of NADH. The acetyl group is attached to a large carrier compound called coenzyme A.
• After the transition step, coenzyme A transports the two-carbon acetyl to the Krebs cycle, where the two carbons enter the cycle. Per turn of the cycle, one acetyl group derived from glycolysis is further oxidized, producing three NADH molecules, one FADH₂, and one ATP by substrate-level phosphorylation, and releasing two CO₂molecules.
• The Krebs cycle may be used for other purposes. Many of the intermediates are used to synthesize important cellular molecules, including amino acids, chlorophylls, fatty acids, and nucleotides.
• Most ATP generated during the cellular respiration of glucose is made by oxidative phosphorylation.
• An electron transport system (ETS) is composed of a series of membrane-associated protein complexes and associated mobile accessory electron carriers. The ETS is embedded in the cytoplasmic membrane of prokaryotes and the inner mitochondrial membrane of eukaryotes.
• Each ETS complex has a different redox potential, and electrons move from electron carriers with more negative redox potential to those with more positive redox potential.
• To carry out aerobic respiration, a cell requires oxygen as the final electron acceptor. A cell also needs a complete Krebs cycle, an appropriate cytochrome oxidase, and oxygen detoxification enzymes to prevent the harmful effects of oxygen radicals produced during aerobic respiration.
• Organisms performing anaerobic respiration use alternative electron transport system carriers for the ultimate transfer of electrons to the final non-oxygen electron acceptors.
• Microbes show great variation in the composition of their electron transport systems, which can be used for diagnostic purposes to help identify certain pathogens.
• As electrons are passed from NADH and FADH₂ through an ETS, the electron loses energy. This energy is stored through the pumping of H⁺ across the membrane, generating a proton motive force.
• The energy of this proton motive force can be harnessed by allowing hydrogen ions to diffuse back through the membrane by chemiosmosis using ATP synthase. As hydrogen ions diffuse through down their electrochemical gradient, components of ATP synthase spin, making ATP from ADP and P_i by oxidative phosphorylation.
• Aerobic respiration forms more ATP (a maximum of 34 ATP molecules) during oxidative phosphorylation than does anaerobic respiration (between one and 32 ATP molecules).

Contributors and Attributions

• Nina Parker, (Shenandoah University), Mark Schneegurt (Wichita State University), Anh-Hue Thi Tu (Georgia Southwestern State University), Philip Lister (Central New Mexico Community College), and Brian M. Forster (Saint Joseph’s University) with many contributing authors. Original content via Openstax (CC BY 4.0; Access for free at https://openstax.org/books/microbiology/pages/1-introduction)