Download 2/1/12 Metabolism

4.1 Nutrition and Cell Chemistry
• Metabolism
– The sum total of all chemical reactions that occur
in a cell
•
Catabolic reactions (catabolism)
– Energy-releasing metabolic reactions
• Anabolic reactions (anabolism)
– Energy-requiring metabolic reactions
• Most knowledge of microbial metabolism is
based on study of laboratory cultures
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4.1 Nutrition and Cell Chemistry
• Nutrients
– Supply of monomers (or precursors of)
required by cells for growth
• Macronutrients
– Nutrients required in large amounts
• Micronutrients
– Nutrients required in trace amount
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4.1 Nutrition and Cell Chemistry
• Carbon
–
–
–
–
–
Required by all cells
Typical bacterial cell ~50% carbon (by dry weight)
Major element in all classes of macromolecules
Heterotrophs use organic carbon
Autotrophs use inorganic carbon
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4.1 Nutrition and Cell Chemistry
• Nitrogen
– Typical bacterial cell ~12% nitrogen
(by dry weight)
– Key element in proteins, nucleic acids, and
many more cell constituents
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4.1 Nutrition and Cell Chemistry
• Other Macronutrients
– Phosphorus (P)
• Synthesis of nucleic acids and phospholipids
– Sulfur (S)
• Sulfur-containing amino acids (cysteine and
methionine)
• Vitamins (e.g., thiamine, biotin, lipoic acid) and
coenzyme A
– Potassium (K)
• Required by enzymes for activity
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4.1 Nutrition and Cell Chemistry
• Other Macronutrients (cont’d)
– Magnesium (Mg)
• Stabilizes ribosomes, membranes, and
nucleic acids
• Also required for many enzymes
– Calcium (Ca)
• Helps stabilize cell walls in microbes
• Plays key role in heat stability of endospores
– Sodium (Na)
• Required by some microbes (e.g., marine
microbes)
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4.1 Nutrition and Cell Chemistry
• Iron
– Key component of cytochromes and FeS
proteins involved in electron transport
– Under anoxic conditions, generally ferrous
(Fe2+) form; soluble
– Under oxic conditions: generally ferric
(Fe3+) form; exists as insoluble minerals
– Cells produce siderophores (iron-binding
agents) to obtain iron from insoluble mineral
form (Figure 4.2)
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4.2 Culture Media
• Culture Media
– Nutrient solutions used to grow microbes in
the laboratory
• Two broad classes
– Defined media: precise chemical composition
is known
– Complex media: composed of digests of
chemically undefined substances (e.g., yeast
and meat extracts)
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4.2 Culture Media
• Selective Media
– Contains compounds that selectively inhibit
growth of some microbes but not others
• Differential Media
– Contains an indicator, usually a dye, that
detects particular chemical reactions
occurring during growth
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4.5 Catalysis and Enzymes
• Enzymes
–
–
–
–
–
Biological catalysts
Typically proteins (some RNAs)
Highly specific
Generally larger than substrate
Typically rely on weak bonds
• Examples: hydrogen bonds, van der Waals
forces, hydrophobic interactions
– Active site: region of enzyme that binds
substrate
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Figure 4.7
Substrate
Products
Active site
Free lysozyme
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Enzyme-substrate
complex
Free lysozyme
III. Oxidation–Reduction and EnergyRich Compounds
• 4.6 Electron Donors and Electron Acceptors
• 4.7 Energy-Rich Compounds and Energy
Storage
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4.6 Electron Donors and Electron
Acceptors
• Energy from oxidation–reduction (redox)
reactions is used in synthesis of energy-rich
compounds (e.g., ATP)
• Redox reactions occur in pairs (two half
reactions; Figure 4.8)
• Electron donor: the substance oxidized in a
redox reaction
• Electron acceptor: the substance reduced in a
redox reaction
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Essentials of Catabolism
•
•
•
•
•
4.8 Glycolysis
4.9 Respiration and Electron Carriers
4.10 The Proton Motive Force
4.11 The Citric Acid Cycle
4.12 Catabolic Diversity
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4.8 Glycolysis
• Two reaction series are linked to energy
conservation in chemoorganotrophs:
fermentation and respiration (Figure 4.13)
• Differ in mechanism of ATP synthesis
– Fermentation: substrate-level phosphorylation;
ATP directly synthesized from an energy-rich
intermediate
– Respiration: oxidative phosphorylation; ATP
produced from proton motive force formed by
transport of electrons
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4.8 Glycolysis
• Fermented substance is both an electron donor
and an electron acceptor
• Glycolysis (Embden-Meyerhof pathway): a
common pathway for catabolism of glucose
(Figure 4.14)
– Anaerobic process
– Three stages
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Figure 4.14
Stage I
Glucose
Stage II
Pyruvate
2 lactate
Stage III
2 Pyruvate
2 ethanol  2 CO2
Intermediates
1,3-Bisphosphoglycerate
Enzymes
Phosphoglycerokinase
Glucose 6-P
3-P-Glycerate
Hexokinase
Phosphoglyceromutase
Fructose 6-P
2-P-Glycerate
Isomerase
Enolase
Fructose 1,6-P
Phosphoenolpyruvate
Phosphofructokinase
Pyruvate kinase
Dihydroxyacetone-P
Aldolase
Lactate dehydrogenase
Glyceraldehyde-3-P
Triosephosphate isomerase
Pyruvate decarboxylase
Glyceraldehyde-3-P
dehydrogenase
Alcohol dehydrogenase
Energetics
Yeast
Lactic acid bacteria
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4.8 Glycolysis
• Glycolysis
– Glucose consumed
– Two ATPs produced
– Fermentation products generated
• Some harnessed by humans for consumption
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4.9 Respiration and Electron Carriers
• Aerobic Respiration
– Oxidation using O2 as the terminal electron
acceptor
– Higher ATP yield than fermentations
• ATP produced at the expense of the proton
motive force, which is generated by electron
transport
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4.9 Respiration and Electron Carriers
• Electron Transport Systems
– Membrane associated
– Mediate transfer of electrons
– Conserve some of the energy released during
transfer and use it to synthesize ATP
– Many oxidation–reduction enzymes are involved in
electron transport (e.g., NADH dehydrogenases,
flavoproteins, iron–sulfur proteins, cytochromes)
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4.9 Respiration and Electron Carriers
• NADH dehydrogenases: proteins bound to
inside surface of cytoplasmic membrane;
active site binds NADH and accepts 2
electrons and 2 protons that are passed to
flavoproteins
• Flavoproteins: contains flavin prosthetic
group (e.g., FMN, FAD) that accepts 2
electrons and 2 protons but only donates the
electrons to the next protein in the chain
(Figure 4.15)
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Figure 4.15
Isoalloxazine ring
Ribitol
Oxidized
Reduced
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4.9 Respiration and Electron Carriers
• Cytochromes
– Proteins that contain heme prosthetic groups
(Figure 4.16)
– Accept and donate a single electron via the
iron atom in heme
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Figure 4.16
Porphyrin
ring
Pyrrole
Heme (a porphyrin)
Protein
Histidine-N
N-Histidine
Cysteine-S
S-Cysteine
Amino acid Amino acid
Cytochrome
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4.9 Respiration and Electron Carriers
• Iron–Sulfur Proteins
– Contain clusters of iron and sulfur (Figure 4.17)
• Example: ferredoxin
– Reduction potentials vary depending on number
and position of Fe and S atoms
– Carry electrons
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Figure 4.17
Cysteine
Cysteine
Cysteine
Cysteine
Cysteine
Cysteine
Cysteine
Cysteine
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4.9 Respiration and Electron Carriers
• Quinones
– Hydrophobic non-protein-containing
molecules that participate in electron
transport (Figure 4.18)
– Accept electrons and protons but pass along
electrons only
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4.10 The Proton Motive Force
• Electron transport system oriented in
cytoplasmic membrane so that electrons are
separated from protons (Figure 4.19)
• Electron carriers arranged in membrane in order
of their reduction potential
• The final carrier in the chain donates the
electrons and protons to the terminal electron
acceptor
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Figure 4.19
E0(V)
0.22
ENVIRONMENT
0.0
Complex II
Q
cycle
Fumarate
0.1
CYTOPLASM
0.36
0.39
E0(V)
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Succinate
4.10 The Proton Motive Force
• During electron transfer, several protons are
released on outside of the membrane
– Protons originate from NADH and the
dissociation of water
• Results in generation of pH gradient and an
electrochemical potential across the
membrane (the proton motive force)
– The inside becomes electrically negative and
alkaline
– The outside becomes electrically positive and
acidic
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4.10 The Proton Motive Force
• Complex I (NADH:quinone oxidoreductase)
– NADH donates e to FAD
– FADH donates e to quinone
• Complex II (succinate dehydrogenase complex)
– Bypasses Complex I
– Feeds e and H+ from FADH directly to
quinone pool
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4.10 The Proton Motive Force
• Complex III (cytochrome bc1 complex)
– Transfers e from quinones to cytochrome c
– Cytochrome c shuttles e to cytochromes a
and a3
• Complex IV (cytochromes a and a3)
– Terminal oxidase; reduces O2 to H2O
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4.10 The Proton Motive Force
• ATP synthase (ATPase): complex that converts
proton motive force into ATP; two components
(Figure 4.20)
– F1: multiprotein extramembrane complex, faces
cytoplasm
– Fo: proton-conducting intramembrane channel
– Reversible; dissipates proton motive force
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Figure 4.20









F1
F1
In
In
b2



c
a
Membrane
c12
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b2

Fo
a
Fo
Out
Out
4.11 The Citric Acid Cycle
• Citric acid cycle (CAC): pathway through
which pyruvate is completely oxidized to CO2
(Figure 4.21a)
– Initial steps (glucose to pyruvate) same as
glycolysis
– Per glucose molecule, 6 CO2 molecules
released and NADH and FADH generated
– Plays a key role in catabolism and
biosynthesis
• Energetics advantage to aerobic respiration
(Figure 4.21b)
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Figure 4.21a
Pyruvate (three carbons)
C2
C4
Acetyl-CoA
C5
C6
Oxalacetate2
Citrate3
Aconitate3
Malate2
Isocitrate3
Fumarate2
Succinate2
-Ketoglutarate2
Succinyl-CoA
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Figure 4.21b
Energetics Balance Sheet for Aerobic Respiration
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4.11 The Citric Acid Cycle
• The citric acid cycle generates many compounds
available for biosynthetic purposes
 -Ketoglutarate and oxalacetate (OAA):
precursors of several amino acids; OAA also
converted to phosphoenolpyruvate, a precursor
of glucose
– Succinyl-CoA: required for synthesis of
cytochromes, chlorophyll, and other tetrapyrrole
compounds
– Acetyl-CoA: necessary for fatty acid biosynthesis
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4.12 Catabolic Diversity
• Microorganisms demonstrate a wide range of
mechanisms for generating energy (Figure 4.22)
–
–
–
–
–
Fermentation
Aerobic respiration
Anaerobic respiration
Chemolithotrophy
Phototrophy
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Figure 4.22
Fermentation
Carbon flow
Organic compound
Carbon flow in
respirations
Electron transport/
generation of pmf
Biosynthesis
Aerobic respiration
Chemotrophs
Electron
acceptors
Anaerobic respiration
Chemoorganotrophy
Electron transport/
generation of pmf
Electron
acceptors
Biosynthesis
Aerobic respiration
Anaerobic respiration
Chemolithotrophy
Light
Photoheterotrophy
Phototrophs
Organic
compound
Photoautotrophy
Electron
transport
e
donor
Generation of pmf
and reducing power
Biosynthesis
Phototrophy
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Biosynthesis
4.12 Catabolic Diversity
• Anaerobic Respiration
– The use of electron acceptors other than
oxygen
• Examples include nitrate (NO3), ferric iron
(Fe3+), sulfate (SO42), carbonate (CO32),
certain organic compounds
– Less energy released compared to aerobic
respiration
– Dependent on electron transport, generation
of a proton motive force, and ATPase activity
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4.12 Catabolic Diversity
• Chemolithotrophy
– Uses inorganic chemicals as electron donors
• Examples include hydrogen sulfide (H2S), hydrogen
gas (H2), ferrous iron (Fe2+), ammonia (NH3)
– Typically aerobic
– Begins with oxidation of inorganic electron donor
– Uses electron transport chain and proton motive
force
– Autotrophic; uses CO2 as carbon source
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4.12 Catabolic Diversity
• Phototrophy: uses light as energy source
– Photophosphorylation: light-mediated ATP
synthesis
– Photoautotrophs: use ATP for assimilation of
CO2 for biosynthesis
– Photoheterotrophs: use ATP for assimilation
of organic carbon for biosynthesis
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