MICROM 301: Week 3 Metabolism

Independent Study Guide:

I. Principles of Metabolism (section 6.1) Cells must: (figure 6.1) • Synthesize new components (anabolism/biosynthesis) • Harvest energy and convert it to a usable form (catabolism)

Harvesting energy: Exergonic vs endergonic reactions Components of metabolic pathways (figure 6.4)

Starting compound à Intermediate à End product Pathways - linear, branched, cyclical

Enzymes (figure 6.5)

Substrate ® Product A specific, unique enzyme catalyzes each biochemical reaction (complete metabolic pathways are in Appendix III)

enzyme-substrate complex, active site (figure 6.11) Cofactors (figure 6.12)

act in conjunction with certain enzymes coenzymes are organic cofactors (table 6.4)

Environmental factors that influence enzyme activity (figure 6.13) Role of temperature, pH

The role of ATP (figure 2.29 and 6.6) • Substrate level phosphorylation • Oxidative phosphorylation • Photophosphorylation

The role of the chemical energy source

Oxidation/reduction reactions (redox reactions) (figure 6.8) OIL – oxidation is loss; RIG – reduction is gain Protons often follow electrons (i.e. a hydrogen atom is extracted/added)

General rules

• If a compound gains oxygen or loses hydrogen, the reaction is an oxidation • If a compound loses oxygen or gains hydrogen, the reaction is a reduction

The role of electron carriers ("reducing power")

Electron carriers (see table 6.1) NAD+, FAD, NADP+

Precursor metabolites – intermediates of catabolism also used in biosynthesis (table 6.2; know concept but DO NOT MEMORIZE)

II. Review Redox reactions

III. Using Precursor Metabolites in Biosynthesis (figure 6.28)

IV. Catabolism of Organic Compounds Other than Glucose (section 6.6)

Compounds enter the central metabolic pathways as precursor metabolites (figure 6.24) Independent Study Questions: 1. What is the difference between anabolism and catabolism? 2. Would a reaction that releases energy be exergonic or endergonic? 3. What is an enzyme? What factors influence enzymatic activity? 4. What about the chemical structure of ATP makes its phosphate bonds "high energy"? 5. How is substrate-level phosphorylation different from oxidative phosphorylation? 6. Why must oxidation and reduction reactions always be coupled? 7. Is removal of a hydrogen atom generally an oxidation or a reduction? 8. What is a precursor metabolite? 9. Where do the electrons carried by NADH enter the electron transport chain relative to

those carried by FADH2? 10. Which is the only pathway that provides a precursor metabolite for nucleotide

biosynthesis? 11. What parts of the central metabolic pathway provide precursor metabolites for lipid

synthesis? 12. How would a bacterium use protein as an energy source? 13. How would a bacterium use lipids as an energy source?

No need to memorize which exact precursor metabolite makes which specific biomolecule and how that happens. Just know which pathway contributes to their synthesis. Example, LPS comes from a precursor metabolite made in glycolysis.

Video Outlines I. Principles of Metabolism (review of independent study material)

Overview of Metabolic pathways

The role of enzymes Allosteric regulation (figure 6.14) Allosteric site vs. active site

Enzyme inhibition (table 6.5)

Competitive inhibition Sulfa drugs as an example (figure 6.15)

Non-competitive inhibition Regulation (allosteric)

The role of ATP (figure 2.29 and 6.6)

• Substrate level phosphorylation - chemical energy is harvested

• Oxidative phosphorylation – the energy of proton motive force (PMF) is harvested; chemical energy is used to create the PMF (involves an electron transport chain)

• Photophosphorylation – the energy of proton motive force is harvested;

radiant energy is used to create the proton motive force (involves an electron transport chain)

The role of electron carriers ("reducing power")

As glucose is oxidized to CO2 (glucose + 6 O2 ® 6 CO2 + 12 H2O), 12 pairs of electrons are removed from glucose (snatched by electron carriers) • Passed to the electron transport chain (used to create the proton motive

force); ultimately passed to a terminal electron acceptor (see figure 3.26) • Used in biosynthesis (to reduce compounds)

II. Overview of Catabolism Glucose + O2 (C6H12O6) Carbon dioxide + H2O

Energy

Energy

Energy

ATP (substrate-level phosphorylation) NADH/FADH2 ® electron transport chain ¯ proton motive force

¯ ATP (oxidative phosphorylation)

Energy source Terminal electron acceptor

Glycolysis Pentose phosphate pathway Transition step TCA cycle Respiration Fermentation

FIGURE 6.10

III. Central Metabolic Pathways (section 6.3)

TCA cycle (aka Kreb's cycle, citric acid cycle (FIGURE 6.17)

Acetyl CoA (2 C) ® 2 CO2 (twice per glucose)

• ATP • 3 NADH • FADH2 • 2 different precursor

metabolites

Glycolysis (figure 6.16) Glucose ® 2 pyruvate

• 2 ATP (net gain) • 2 spent, 4 made • 2 NADH • six different

precursor metabolites

Pentose phosphate pathway (not illustrated) Glucose ® intermediate of glycolysis

• NADPH (amount varies)

• two different precursor metabolites

Transition step (FIGURE 6.17) Pyruvate (3 C) ® acetyl CoA (2 C) + CO2 (twice per glucose)

• NADH • 1 precursor

metabolite

IV. Cellular Respiration (section 6.4) The central metabolic pathways oxidize glucose …but where do the electrons go????? Electron transport chain of mitochondria (part of figure 6.19) Electron transport chain of E. coli (figure 6.20) Aerobic respiration Anaerobic respiration

o NO3 is the TEA Overall maximum energy yield of glucose metabolism using aerobic respiration (ignoring the pentose phosphate pathway) (Figure 6.21) 4 ATP - substrate level phosphorylation 10 NADH 2 FADH2

Electron transport chain

®oxidative phosphorylation

}

Energy Source vs Terminal Electron Acceptor (figure 6.7)

Glucose + 6 O2 ® 6 CO2 + 12 H2O

Application: Microbiology of Wastewater treatment (Section 29.1)

BOD (biochemical oxygen demand): amount of O2 required for microbial decomposition of organic matter in a sample

ATP Synthase – Using the Proton Motive Force to Synthesize ATP

harvesting proton motive force ATP production Flagellum rotation Active transport

V. Fermentation (section 6.5, part of

figure 6.10)

• Used by some cells when respiration is not an option – lack of TEA, or no electron transport chain

• Oxidation of glucose stops at pyruvate

• Passes electrons from NADH to pyruvate or a derivative

End products of fermentation (figure 6.23) VI. Review (Table 6.3) Review/Discussion Questions: 1. What is the mechanism of action of sulfa drugs? 2. Mercury kills bacteria (and other cells) because it is a non-competitive enzyme inhibitor.

What does this mean? 3. What is the fate of the electrons carried by NADH and FADH2? What about NADPH? 4. What are the central metabolic pathways? Why are they called central? 5. What is the endpoint of glycolysis? of the TCA cycle? 6. What is the transition step? 7. Why does the transition step and TCA cycle have to repeat “twice per glucose molecule”? 8. Identify which central metabolic pathway:

a. can initiate the breakdown of glucose? b. is generally used exclusively for biosynthesis? c. generates the most reducing power? d. generates the most ATP directly? e. Generates the most ATP indirectly? f. includes steps that use ATP?

9. What is proton motive force used for in mitochondria? in bacteria? 10. In the breakdown of glucose, which generates the most ATP: substrate-level

phosphorylation or oxidative phosphorylation?

The logic: • Oxidizes NADH, generating NAD for use in further rounds of glucose

breakdown

• Stops short of the transition step and the TCA cycle, which together generated 5X more reducing power

11. How is fermentation different from respiration? 12. How is NAD regenerated in respiration vs. in fermentation? 13. How is aerobic respiration different from anaerobic respiration? 14. When glucose serves as the energy source, which terminal electron acceptor – NO3 or O2 –

results in the release of the most energy? 15. What is BOD? How will adding a substance with a high BOD affect a body of water like

Greenlake? 16. Give three examples of why the end products of fermentation are important. Independent Study – Wastewater Treatment I. Chapter 29 Introduction Terminology you should be able to define:

• BOD • Bioremediation • Primary Treatment • Secondary Treatment • Septic System • Wastewater • Sludge

Videos – Wastewater Treatment II. Wastewater Treatment Steps in the process of sewage-based wastewater treatment

1. Primary Treatment 2. Secondary Treatment 3. Advanced Treatment 4. Anaerobic Sludge

Digestion 5. Disinfection

III. Other Types of

Sanitation Systems Categorized by location, resource recovery, and means of transportation Many places in the world can’t afford to have large municipal, sewered wastewater facilities

Development of decentralized systems are important

Review/Discussion Questions: 1. Where in these processes are potentially pathogenic bacteria killed? 2. Where in these processes are beneficial microbes used? 3. How are some of the byproducts of these processes used? Videos – Methane and Nitrogen Cycles and Microbes Involved I. Methane branch of carbon cycle (28.5)

A. Methanogens produce CH4

Methane (biogas) is produced by reducing CO2 using H2 as electron donor Performed by Archea Not a lot of energy in this pathway Create proton and sodium gradients to make ATP with B. Methanotrophs and methylotrophs use

CH4

Oxidize methane to carbon dioxide

9/4/2018

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This means, DEWATS is led by some principles, which represent the guide frame for designing

This will be the best DEWATS

Conservation / Recycling :

Water, Nutrients,

Energy

Decentralization : Responsibility,

Capacity, Technology

Simplification: Process,

Technology, O&M

DEWATS Principle

Typical combination of DEWATS module

Septic tank

Anaerobic Baffled Reactor

Anaerobic Filter

Planted Gravel Filter

Pond Systems

Biogas settler

Screening and grit chamber,

Greese/oil trap

Right Combination of DEWATS modules makes efficient/successful treatment system.

II. Nitrogen Cycle

1. Nitrification oxidizes ammonium to nitrite or nitrate

2. Denitrification reduces nitrate to nitrite (anaerobic respiration), and nitrite is usually reduced further to nitrogen gas or ammonium

Nitrogen gas can’t be used by most organisms. Leads to loss of nitrogen from life cycles.

3. Nitrogen Fixation allows for recovery of nitrogen by converting it to ammonium Only microbes can do this at a normal temperatures and pressures. Rhizobia live as endosymbionts with specific types of plants, usually legumes.

Review/Discussion Questions: 1. What category of bacteria produce methane? What would the classification of these

organisms be? 2. What category of bacteria oxidize methane? 3. Distinguish between nitrification, denitrification, and nitrogen fixing. 4. What is the enzyme that in associated with nitrogen fixing? What molecule is this enzyme

sensitive to? Independent Study - Food Microbiology I. Principles of Food Microbiology (CHAP 30 INTRO) II. Factors Influencing the Growth of Microorganisms in Foods (SECTION 30.1)

In general, bacteria grow faster than yeasts, which grow faster than molds

intrinsic factors water availability (aw) - do NOT memorize exact numbers

pH

nutrients biological barriers antimicrobial chemicals

extrinsic factors

temperature atmosphere

IV. Food Preservation (preventing/delaying spoilage) (SECTION 30.5)

kill organisms heat

canning (commercially sterile) pasteurization cooking

pressure irradiation inhibit growth of organisms refrigerate freeze dry/reduce the aw lower the pH add antimicrobial chemicals

Independent Study Questions

1. What is the difference between a fermented food and a spoiled food? 2. Which typically grows fastest - yeasts, bacteria, or molds? 3. Describe 4 intrinsic factors that influence the growth of microorganisms in food. 4. Describe 2 extrinsic factors that influence the growth of microorganisms in food. 5. Which would predominate on improperly stored ham - a Pseudomonas species or a

Staphylococcus species. Explain. 6. How would storage temperature influence the rate of microbial growth? 7. How would storage temperature influence the types of microorganisms that would predominate

on a food product? 8. How is a canned food product different from a pasteurized food product?

dH2O (1.0) Ham

(0.91) Jam (0.85)

Cake (0.7) Fresh meat

(0.99)

Pseudomonas (0.97)

yeasts (0.88) Staphylococcus (0.86)

Milk (6.4)

Spinach (5.6)

Lemons (2.2)

Pseudomonas (5.5)

Lactic acid bacteria (3.5) Yeasts (1.5)

Molds (0.0)

Cherries (3.8)

9. Describe the difference between pasteurization and canning. 10. Describe four general methods to inhibit bacterial growth in a food. 11. Describe two general methods to decrease the aw of a food product.

Videos - Food Microbiology I. Microorganisms in Food and Beverage Production (SECTION 30.2) Desirable change ¬ control conditions Lactic acid fermentations by the lactic acid bacteria glucose ® pyruvate ® lactic acid

Alcoholic fermentations by yeast (Saccharomyces) Glucose ® pyruvate ® ethanol + CO2

Mold growth

Other fermented food products Coffee Chocolate II. Additional Information History of fermented foods Human transition from hunter-gather society to agrarian society coincided with fermentation of grains Review Questions

1. Name three distinct categories of foods that are made using lactic acid bacteria, and an example of a product in each category.

2. Name two genera that fit into the category of "lactic acid bacteria". 3. Name two categories of foods (or beverages) produced using Saccharomyces cerevisiae. 4. Name a food produced using a mold. 5. Name the components of a meal you would serve that uses ONLY fermented foods (or

beverages).