Chapter 5 Microbial Metabolism PDF

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Online Prof. D Hemmerling
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Chapter 5: Microbial Metabolism Objectives Upon completion of this section, you will be able to: 1. Explain the composition and function of enzymes. 2. Describe and give examples of catabolism, anabolism, respiration, and fermentation. 3. Understand the function of the components of respiration and fermentation. 4. Understand the purpose of fermentation.

Metabolism- There are 2 types of metabolic processes 

Metabolism that releases energy is called catabolism. Catabolism results in large macromolecules being broken down into their smaller components. These smaller parts may be burned for energy or they may be used as building blocks to make new macromolecules for the cell.

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Metabolism that consumes energy (uses ATP) is called anabolism. Anabolic processes use energy and simple building blocks to make new macromolecules for the cell.

Enzymes Enzymes are "biological catalysts". It is a component of a chemical reaction which enhances the rate of the reaction but remains unaltered by the process. A cell’s metabolic pathways are regulated by its enzymes. Almost every step in the complex metabolic pathways of the cell is controlled by an enzyme. Enzymes are proteins. All proteins are composed of 20 different amino acids arranged in a linear chain. The 20 amino acids differ in their side chains, which are called R groups. The R groups determine the unique chemical nature of each amino acid. Amino acids fall into four chemical types:    

hydrophobic basic acidic polar non-ionizable.

In a protein, the amino acids are joined by peptide covalent bonds. Enzymes are the tools of life and each enzyme has a specific job description. The chemical nature of an enzyme and therefore its function are determined by the sum of the chemical characteristics of its amino acids and by their arrangements. Every enzyme is unique and has only one job to do. Rules for Enzyme Function: 1. Enzymes are large compared to the molecules they work on. 2. Enzymes work very fast and can carry out reactions thousands of times per second. 3. Each enzyme has a functional structure. This structure gives an enzyme the ability to bind the molecules it is designed to work on. The molecule that an enzymes acts on is called the substrate and the region of the enzyme that interacts with the substrate is called the active site. 4. Enzymes are very specific for particular reactions because they interact specifically with their substrates. Enzymes have this specificity because the three-dimensional shape of the active site fits the substrate both in physical dimensions and in charge interactions. Due to the size difference between enzyme and substrate, the substrate always fits into the enzyme’s active site, which is a fold or groove in the enzyme. The specificity of the enzyme substrate interaction is so high that the change in the location of a single atom in a substrate will render that substrate nonfunctional.

Mechanism of Enzyme Action: 1. The substrate contacts the active site on the enzyme to form an enzyme-substrate complex. 2. The substrate is then transformed into products. 3. These products are released and the enzyme is recovered unchanged.

Cofactors Many enzymes require small molecules called cofactors in order to function correctly. Cofactors can be: 

organic molecules such as vitamins, these are called coenzymes

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inorganic molecules

Cofactors often function by carrying pieces of molecules from one place to another, cofactors are part of the active site (although they can enter and leave) and therefore contact the substrate. This puts them in position to receive a piece removed from the substrate. Two important coenzymes are nicotinamide adenine dinucleotide (NAD+) and nicotinamide adenine dinucleotide phosphate (NADP+). These are electron carriers and are very important in energy metabolism. Whereas NAD+ is primarily involved in catabolic (energy-yielding) reactions, NADP+ is primarily involved in anabolic (energy-requiring) reactions.

Factors Influencing Enzyme Activity 

Temperature. The rate of most chemical reactions increases as the temperature increases but in enzymecatalyzed reactions the enzyme usually works best at one temperature. Therefore, increases in the temperature are detrimental as they destroy cellular proteins. The crucial function of enzymes is to speed up biochemical reactions without significantly raising the temperature. If the temperature even a few degrees above the optimum, the enzyme will denature.

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Substrate concentration. The more substrate that is available (up to a certain point), the better the enzyme works.

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Inhibitors. Enzyme inhibitors are either competitive or noncompetitive. o

Competitive inhibitors chemically mimic the true substrate but do not undergo any sort of reaction to form a product. They are so similar that they fit and bind to the active sites of an enzyme and decrease the amount of binding of the substrate can do because the active site is already filled. Competitive inhibitors are also referred to as substrate analogs. Example of a competitive inhibitor—Sulfanilamide (Sulfa Drug)

o Noncompetitive inhibitors do not compete with the substrate for the enzyme active site. Some bind to other parts of the enzyme and in doing so change the shape of its active site.

Carbohydrate Catabolism- breakdown into smaller parts, releases energy in the process Basic Concepts 

An atom becomes more reduced (undergoes reduction) when it undergoes a chemical reaction in which it gains electrons

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An atom becomes more oxidized when it undergoes a chemical reaction in which it loses electrons. The reaction usually produces energy

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Oxidation and reduction reactions are always coupled: each time one substance is oxidized, another is simultaneously reduced. The pairing of these reactions is called oxidation-reduction or a redox reaction. In metabolic pathways, we are often concerned with the oxidation or reduction of carbon. The thing that gets oxidized is called the electron donor, and the thing that gets reduced is called the electron acceptor.

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Reduced forms of carbon (e.g. hydrocarbons, methane, fats, carbohydrates, alcohols) carry a lot of potential chemical energy stored in their bonds.

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Oxidized forms of carbon (e.g. ketones, aldehydes, carboxylic acids, carbon dioxide) carry very little potential chemical energy in their bonds.

Enzymatic Pathways for Metabolism 

Metabolic reactions take place in a step-wise fashion in which the atoms of the raw materials are rearranged, often one at a time, until the formation of the final product takes place.

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Each step requires its own enzyme.

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The sequence of enzymatically-catalyzed steps from a starting raw material to final end products is called an enzymatic pathway (or metabolic pathway)

Basic Concepts 

Need cofactors for Redox Reactions

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Enzymes that catalyze redox reactions typically require a cofactor to “shuttle” electrons from one part of the metabolic pathway to another part.

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There are two main redox cofactors: NAD and FAD. These are (relatively) small organic molecules in which part of the structure can either be reduced (e.g., accept a pair of electrons) or oxidized (e.g., donate a pair of electrons) electrons have a negative charge

The Cofactors for Redox Reactions when a metabolite is oxidized, NAD accepts 2 electrons and 1 H+ to form NADH(reduced) FAD(oxidized), FAD accepts 2 electrons and 1 H+ to form FADH(reduced) Remember this . . . . . . . NAD and FAD are present only in small (catalytic) amounts – they cannot serve as the final electron acceptor, but must be regenerated (reoxidized) in order for metabolism to continue 

ATP stands for adenosine triphosphate. Remember that a molecule of ATP consists of an adenine, a ribose, and three phosphate groups linked in a small chain.

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When the terminal phosphate group is split from ATP, adenosine diphosphate (ADP) is formed, and energy is released to drive anabolic reactions

The last phosphate in the chain can be removed by hydrolysis (Usually hydrolysis is a chemical process in which a molecule of water is added to a substance) in this case, the ATP becomes ADP, or adenosine diphosphate). To produce energy from glucose, microorganisms use two general processes: Cellular respiration and fermentation. Both start with glycolysis (glyco=sugar, lysis=breakdown) For example, the sugar sucrose may undergo hydrolysis to break into its component sugars, glucose and fructose.

Cellular Respiration Overview 

Heterotrophs – an organism that can not create its own energy, relies on obtaining carbon from organic compounds

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Heterotrophs generate energy by cellular respiration.

Cellular Respiration consists of a series of pathways: 1. Glycolysis [in the cytoplasm] 2. Kreb's cycle or TCA cycle [in the mitochondrial matrix] 3. Electron Transport Chain & Oxidative Phosphorylation [inner mito. MB] #1 Glycolysis -the most common pathway among cell types The net result of glycolysis is to convert a glucose molecule into 2 pyruvic acid molecules with the production of 2 NADH and 2 ATP.   

an anerobic process (does not need oxygen, however the TCA cycle does) Actually a 10 steps process, enzyme-catalyzed which releases energy in small amounts. (we do not need to know all 10 steps) Serves as the beginning for aerobic or anaerobic metabolism.

Aerobic Respiration – occurs to carry the pyruvate onward into TCA cycle and ET chain only if oxygen is present, to produce more ATP 

In eukaryotes, these processes occur in the mitochondria, while in prokaryotes they occur in the cytoplasm.

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If there is no oxygen, cells continue on to fermentation to continue making ATP (see later in chapter)

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The two (3 carbon compound) pyruvic acids enter the mitochondrial matrix (from glycolysis) and is converted into two, 2 carbon compounds called Acetyl CoA (through decarboxylation where it loses 1 molecule of CO2 ) which is then carried in to the Kreb’s Cycle (TCA cycle) REMEMBER FOR EVERY GLUCOSE THAT ENTERS GLYCOLYSIS TWO PYRUVATES ARE PRODUCED

#2 Kreb’s Cycle- also known as the citric acid cycle and the tricarboxylic acid cycle (TCA) 

Each step is catalyzed by a specific enzyme

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CO2 is released as the byproduct of the cycle- this is the CO2 we exhale

NAD+ is an oxidizing agent – it accepts electrons from other molecules and becomes reduced. This reaction forms NADH, which can then be used as a reducing agent to donate electrons. Yields from 2 turns of Krebs (TCA) cycle: 

2 ATP (by substrate-level phosphorylation)

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6 NADH to be used to generate energy by chemiosmosis/Ox. Phos. (Chemiosmosis is the method which cells use to create ATP for energy. ... The electrons move through the electron transport chain to oxygen, where they generate energy which pumps the hydrogen ions against their concentration gradient from matrix to the intermembrane space, so they can flow back down again.)

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2 FADH2 by chemiosmosis/Ox. Phos

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4 CO2

SUMMARY: When the reactions of glycolysis are completed, 2 ATP have been produced, as well as 2 NADH and 2 pyruvate. The pyruvates are transported into the mitochondrion and used to bring new carbons into the Krebs/TCA/citric acid cycle. In the Krebs cycle, 2 ATP, 6 NADH, 2 FADH2, and 4 CO2 have been produced. #3 The Electron Transport Chain- the final step in cellular respiration 

What happens to the NADH and FADH2? ---- who the fuck knows?

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It’s where the electron carriers put their cargo of electrons

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The transfer of electrons from NADH and FADH to the final electron acceptor, molecular oxygen, occurs in an elaborate electron transport chain. The function of this chain is to permit the controlled release of free energy to drive the synthesis of ATP.

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When activated, ATP synthase during the ETC provides a rich yield of 28 more ATP molecules for each glucose molecule processed (from the total 10 NADH and 2 FADH2 molecules formed during glucose processing)

The Two Main functions of the ETC 

Electrons pass down the ETC in a series of redox reactions (An oxidation-reduction (redox) reaction is a type of chemical reaction that involves a transfer of electrons between two atoms where one undergoes oxidation and the other, reduction).

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The electrons entering the electron transport system have a relatively high energy content. As they pass along the chain of electron acceptors, they lose much of their energy, some of which is used to pump protons across the inner mitochondrial membrane.

Oxidative phosphorylation refers to the process by which ATP is synthesized using energy released by electrons as they are transferred to O2.

The TCA and ETC take place in the mitochondria in eukaryotes and in the cytoplasm for prokaryotes (bacteria)

In aerobic respiration among prokaryotes, each molecule of glucose generates 38 ATP molecules: 34 from chemiosmosis plus 4 generated by oxidation in glycolysis and the Krebs cycle. Aerobic respiration among eukaryotes produces a total of only 36 molecules of ATP.

Re-oxidation of NAD+ If something becomes oxidized, it’s losing electrons. Meanwhile, if something is reduced, it is gaining electrons. Electrons = - charge Protons= + charge To oxidize NADH to NAD+, oxidation occurs which means it’s losing electrons. Electrons have a – charge, so to lose electrons would make the + in the NAD During glycolysis, we have to change NADH to NAD+ through re-oxidation. Oxidation happens when a cell loses electrons. Electrons have a – charge, so to lose electrons would make the + in the NAD. In the ETC, the NADH donates it’s H and it floats into the intermembrane space. You’re left with NAD+ which is now positively charged, thus the + Fermentation 1. releases energy from sugars or other organic molecules; 2. does not require oxygen (but can occur in its presence); 3. does not require the use of the Krebs cycle or an electron transport chain; 4. uses an organic molecule synthesized in the cell as the final electron acceptor. 

It can take place in all types of cells.

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Under anaerobic conditions (the absence of oxygen), pyruvic acid can be routed by the organism into one of three pathways: lactic acid fermentation, alcohol fermentation, or cellular (anaerobic) respiration. If there is no oxygen, glucose cannot be broken down during glycolysis. The untapped energy of the glucose remains locked in the pyruvate molecules and is eventually converted to lactate if they do not enter the pathway that eventually leads to oxidative phosphorylation (pathway is the TCA and ETC cycle)

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Fermentation produces only small amounts of ATP (only one or two ATP molecules for each molecule of starting material) because much of the original energy in glucose remains in the chemical bonds of the organic endproducts, such as lactic acid or ethanol. However, the advantage of fermentation for a cell is that it produces ATP quickly.

 It is important to note that for some microorganisms, regardless of oxygen concentrations, fermentation is the only option. 

Need a steady supply of NAD+ to accept electrons.

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Main purpose is to re-oxidize NADH to NAD+ to send them back to glycolysis. NAD+ must be reoxidized so that there is enough NAD+ for glycolysis (it is too energy expensive to just make more NAD+)

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During fermentation, electrons are transferred (along with protons) from reduced coenzymes (NADH, NADPH) to pyruvic acid

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It is a wasteful process because there is still potential (chemical energy) in the waste products

Figure 5.17 A summary of aerobic respiration in prokaryotes. Glucose is broken down completely to carbon dioxide and water, and ATP is generated. This process has three major phases: glycolysis, the Krebs cycle, and the electron transport chain. The preparatory step is between glycolysis and the Krebs cycle. The key event in aerobic respiration is that electrons are picked up from intermediates of glycolysis and the Krebs cycle by NAD+ or FAD and are carried by NADH or FADH2 to the electron transport chain. NADH is also produced during the conversion of pyruvic acid to acetyl CoA. Most of the ATP generated by aerobic respiration is made by the chemiosmotic mechanism during the electron transport chain phase; this is called oxidative

phosphorylation.

WHAT ARE THE INPUTS AND OUTPUTS OF A GLYCOLYIS REACTION?...