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Metabolism
• Metabolism in bacteria is similar to that in eukaryotes.
• Some bacterial enzymes (especially metabolic enzymes, like oxidase) can be interchanged with human enzymes in laboratory experiments.
• However, bacteria also have unique enzymes that allow them to adapt to many niches.
• How do horses and cows digest cellulose?
• How do bacteria live at the depths of the ocean?– They have special enzymes which have been adapted for specific environments.
•Metabolism: pertains to all chemical reactions and physical workings of the cell
•Anabolism:- synthesis of molecules, requires the input of energy
•Catabolism:- breaks the bonds of larger molecules, releases energy
Metabolism
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All Catabolic reactions involve electron transfer
• Electron transfer:– allows energy to be captured in high‐energy bonds in ATP and similar
molceules.
– Directly related to oxidation & reduction (remember those redox reactions from chemistry?)
• Oxidation=loss of electrons
• Reduction=gain of electrons
• Imagine A=organic molecule like glucose
• Imagine B=NAD+ coenzyme, an electron carrier
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.R
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ANABOLISM
ANABOLISM
ANABOLISM
Nutrientsfrom outsideor frominternalpathways
Glycolysis
Krebs cycle
Respiratorychain
Fermentation
Yields energy Uses energy Uses energy Uses energy
Some assemblyreactions occurspontaneously
Complex lipids
RNA + DNA
Peptidoglycan
Proteins
Amino acids
Sugars
Nucleotides
Fatty acidsGlyceraldehyde-3-P
Acetyl CoA
Pyruvate
CATABOLISM
Glu
Glucose
Precursormolecules
Macromolecules
Bacterialcell
Buildingblocks
Simplified Model of Metabolism
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• In order for bonds to be FORMED or BROKEN, there has to be a minimal amount of energy available. This minimal amount of energy is termed “activation energy.”
How do chemical reactions take place? How do chemical reactions take place?
• Activation energy can be in the form of temperature or pressure, etc. to increase the number of particle collisions.
• Paradoxically, the temperature and pressure that humans and bacteria would require for their chemical reactions would KILL them!
• How do we solve this problem?• ENZYMES!!
• Enzymes reduce the amount of activation energy needed for chemical reactions and speed them up so that life can continue.
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Enzymes
• Enzymes have specific active sites that bind to specific substrates.
• Bonds formed between the substrate and enzyme are weak and easily reversible
• Enzyme are fast!
- the number of substrate molecules converted per enzyme per second
- Catalase reacts several million times per second
- lactate dehydrogenase reacts a thousand times per second
Specific active sites arise due to the folding of the protein
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Enzyme‐substrate interactions
• Substrates specifically bind to the active sites on the enzyme
– “lock‐and‐key”
– Induced fit
• Once the reaction is complete, the product is released and the enzyme reused
Lock-and-key model
Induced fit model
Enzyme‐Substrate Reactions
•The need of microorganisms for trace elements arises from their roles as cofactors for enzymes
- iron, copper, magnesium, manganese, zinc, cobalt, selenium, etc.
•Participate in precise functions between the enzyme and substrate
- help bring the active site and substrate close together
- participate directly in chemical reactions with the enzyme‐substrate complex
Cofactors
Metalliccofactor
- organic compounds that work in conjunction with an enzyme
- general function is to remove a chemical group from one substrate molecule and add it to another substrate molecule
- carry and transfer hydrogen atoms, electrons, carbon dioxide, and amino groups
- many derived from vitamins
Coenzymes
Coenzyme
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Apoenzyme
• The main enzyme portion is a globular protein called an apoenzyme
Example of how a coenzyme transfers chemical groups from one substrate to another
Constitutive enzymes:always present in relatively constant amounts regardless of the amount of substrate
Regulation of Enzymes
Regulated enzymes:production is turned on (induced) or turned off (repressed) in responses to changes in concentration of the substrate
Regulation of Enzymes
One type of genetic control of enzyme synthesis
Regulation of Enzymes
•If E. coli is inoculated with only lactose, it will produce the enzyme lactase to hydrolyze the lactose into glucose and galactose
•If E. coli is inoculated with only sucrose, it will cease to synthesizing lactase and begin synthesizing sucrase
•Benefits:• Allows the organism to utilize a variety of nutrients
• Prevents wasting energy by making enzymes for a substrate that is not present
Enzyme Induction in E. coli
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•Activity of enzymes influenced by the cell’s environment• Natural temperature, pH, osmotic pressure• Denaturation: weak bonds that maintain the native shape of
the enzyme are broken
Regulation of Enzyme Function
Competitive inhibition- inhibits enzyme activity
by supplying a molecule that resembles the enzyme’s normal substrate
- “mimic” occupies the active site, preventing the actual substrate from binding
Inhibition of Enzymes
Noncompetitive inhibition
Noncompetitive inhibitors bind to an “allosteric” or “other” site on the enzyme, not the active site.
Inhibition of Enzymes
•Often occur in a multistep series or pathway, with each step catalyzed by an enzyme
•Product of one reaction is often the reactant (substrate) for the next, forming a linear chain or reaction
Metabolic Pathways
A
B
C
D
E
Linear
Example:Glycolysis
Many pathways have braches that provide alternate methods for nutrient processing
Metabolic Pathways
O2
O
O1
M
N
P
Q
R
M
A
B
C
N
X
Y
Z
Branched
Convergent
Example:Amino acidsynthesis
Divergent Other pathways have a cyclic form, in which the starting molecule is regenerated to initiate another turn of the cycle
Metabolic Pathways
U
V
W
X
Z
Y
Cyclic
T input
KrebsCycle
S product
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Metabolic pathways do not stand alone; they are interconnected and merge at many sites
Metabolic Pathways
CA
TA
BO
LIS
MA
NA
BO
LIS
M
Gly
co
lys
is
Beta oxidationDeamination
GLUCOSE
Metabolicpathways
Simplepathways
Pyruvic acid
Acetyl coenzymeA
KrebsCycle
NH3 H2O
CO2
Building block
Macromolecule
Cellstructure
Membranesstorage
Cell wallstorage
Enzymes/Membranes
Chromosomes
Lipids/Fats
Starch/CelluloseProteins
Nucleicacids
Fatty acidsCarbohydratesAmino acidsNucleotides
•Oxidation: loss of electrons
•Reduction: gain of electrons
•Oxidoreductases: enzymes that remove electrons from one substrate and add them to another
- their coenzyme carriers are nicotinamide adenine dinucleotide (NAD+) and flavin adenine dinucleotide (FAD)
Oxidation and Reduction
-+
Na 2 8 7Cl2 8 1
Oxidizing agentaccepts electrons.
Reducing agentgives up electrons.
Na 2 8 2 8 8Cl
Oxidizedcation
Reducedanion
Electron carriers resemble shuttles that load and unload electrons and hydrogens to facilitate transfer of redox energy
electrons availablein NADH and FADH2
electrons are transferred in many cases along with protons as hydrogen atoms since one hydrogen atom (H) = 1 proton (H+) plus 1 electron (e‐)
H+
P
P
P
P
H++NAD+ NAD H
Reduced Nicotinamide
From substrate
Oxidized Nicotinamide
Adenine
Ribose
NH22H2e:
H
C
C C
C C
O
CH
NH2
H
C
C C
C C
O
C
N N
Electron Carriers: Cell’s Reducing Power
•Three‐part molecule- nitrogen base (adenine)- 5‐carbon sugar (ribose)- chain of three
phosphate groups bonded to ribose
- phosphate groups are bulky and carry negative charges, causing a strain between the last two phosphates making it very volatile
N
NN
N N
H H
H
H
O
HHH H
O
O
O
O
P O
O
H
HPP
Adenine
AdenosineAdenosine
Diphosphate(ADP)
AdenosineTriphosphate
(ATP)
HO
OH OH OH
OH
Ribose
OHBond that releasesenergy when broken
Adenosine Triphosphate‐ ATP
ATP can be used to phosphorylate an organic molecule
Ex. Phosphorylation of glucose to activate its catabolism
ATP and Phosphorylation
Fermentation
ANAEROBIC RESPIRATION FERMENTATION
ATP ATP
AEROBIC RESPIRATION
CO2
NAD H
ATP
CO2
NAD H
ATP
NAD HCO2
ATPFADH2
Using organiccompounds as
electron acceptor
Electron Transport System Electron Transport System
Alcohols, acids
2 ATPs2–36 ATPs36–38 ATPsMaximum net yield
Yields variableamount ofenergy
Yields 2 ATPs
Yields 2 ATPs
CO2
NAD H
ATP
NAD H
FADH2 ATP
CO2
KrebsCycle
KrebsCycle
Using O2 as electron acceptor Using non- O2 compound as electron acceptor
(So42–, NO3–, CO3
2–)
Gly
co
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s
Gly
co
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s
Gly
co
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s
Overview of the Three Main Catabolic Pathways
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- A series of reactions that converts glucose to CO2 and allows the cell to recover significant amounts of energy- Complete breakdown of pyruvic acid into inorganic molecules- Relies on free oxygen as the final electron- Characteristic of many bacteria, fungi, protozoa, and animals
Aerobic Respiration: Overview
ATP
Electron Transport System
36–38 ATPsMaximum net yield
CO2
NAD H
ATP
NAD H
FADH2 ATP
CO2
KrebsCycle
Using O2 as electron acceptor
Gly
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s
- uses NO3‐, SO4
2‐, CO33‐, and other
oxidized compounds as final electron acceptors
- Like aerobic respiration there is a complete breakdown of pyruvic acid into inorganic molecules
- Unlike aerobic respiration, anaerobic respiration does not use all of the steps in the Kreb’s cycle.
- characteristic of bacteria that require or tolerate anaerobic conditions
Anaerobic Respiration: Overview
ATP
CO2
NAD H
ATP
NAD HCO2
ATPFADH2
Electron Transport System
2–36 ATPsMaximum net yield
KrebsCycle
Using non- O2 compound as electron acceptor
(So42–, NO3–, CO3
2–)
Gly
co
lysi
s
• Unlike aerobic and anaerobic respiration pyruvic acid is notcompletely broken down into inorganicmolecules
• Pyruvic acid is partially broken down into organic compounds that are the final electron acceptors. Ex. lactic acid, ethanol…
• oxygen is not required
Fermentation: Overview
Fermentation
CO2
NAD H
ATP
Using organiccompounds as
electron acceptor
Alcohols, acids
2 ATPsMaximum net yield
Gly
co
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s
GlucoseGlucose
Fermentation
ANAEROBIC RESPIRATION FERMENTATION
ATP ATP
AEROBIC RESPIRATION
CO2
NAD H
ATP
CO2
NAD H
ATP
NAD HCO2
ATPFADH2
Using organiccompounds as
electron acceptor
Electron Transport System Electron Transport System
Alcohols, acids
2 ATPs2–36 ATPs36–38 ATPsMaximum net yield
Yields variableamount ofenergy
Yields 2 ATPs
Yields 2 ATPs
CO2
NAD H
ATP
NAD H
FADH2 ATP
CO2
KrebsCycle
KrebsCycle
Using O2 as electron acceptor Using non- O2 compound as electron acceptor
(So42–, NO3–, CO3
2–)
Gly
co
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s
Gly
co
lysi
s
Gly
co
lysi
s
Overview of the Three Main Catabolic Pathways Stop
• https://www.youtube.com/watch?v=‐Gb2EzF_XqA
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• Oxidation of glucose into pyruvate, which yields energy in the pathways that follow
• Occurs in cytoplasm of both eukaryotes and prokaryotes‐Does this go against the Endosymbiotic theory?
Table 7.2
One reaction breaks fructose-1,6-diphosphateinto two 3-carbon molecules.
Five reactions convert each 3 carbon moleculeinto the 3C pyruvate.
Pyruvate is a molecule that is uniquely suited for chemicalreactions that will produce reducing power (which willeventually produce ATP).
C C C C C C
Fructose-1, 6-diphosphate
C C C C C C
C C CC C C
C C CC C C
Glycolysis
Energy Lost or Gained
Uses 2 ATPs
Overview Details
Three reactions alter and rearrange the6-C glucose molecule into 6-C fructose-1,6diphosphate.
Yields 4 ATPs and 2 NADHs
Total Energy Yield: 2 ATPs and2 NADHs
Glucose
Pyruvate Pyruvate
Glycolysis
Table 7.3
Each acetyl CoA yields 1 GTP, 3 NADHs,1 FADH, and 2 CO2 molecules.
Total Yield per 2 acetyl CoAs:CO2: 4 In the course of seven more
reactions, citrate is manipulatedto yield energy and CO2 andoxaloacetate is regenerated.
Intermediate molecules on thewheel can be shunted into othermetabolic pathways as well.
In the first reaction, acetyl CoAdonates 2Cs to the 4C moleculeoxaloacetate to form 6C citrate.
C C C
Energy: 2 GTPs, 6 NADHs, 2 FADHs
Pyruvate
CC CC CC
Details
The Krebs Cycle
Energy Lost or Gained Overview
Pyruvate
The 3C pyruvate is converted to2C acetyl CoA in one reaction.
Otherintermediates GTP
CO2
CO2
Yields:3 NADHs1 FADH2
Citrate
Oxaloacetate
Acetyl CoA
Remember: Thishappens twice for
each glucosemolecule that
enters glycolysis.
One CO2 is liberated and one NADH isformed.
C C C C
C C C C C C
C CC
The Krebs Cycle• Produces 2 ATP for each molecule of glucose• Purpose is to produce NADH and FADH2 to be fed into the ETC• Occurs in the cytoplasm of bacteria and in the mitochondrial matrix
of eukaryotes
Table 7.4 The Respiratory (Electron Transport) Chain
Anaerobicrespirers
Aerobicrespirers
CytoplasmH2O NO2
– HS–
O2
H+
CellmembraneWith ETS
Cell wall
H+
H+
H+
H+
H+
H+
H+
H+
H+
H+
H+H+
H+
H+
Cytochromes
NAD H
ATPADP
ATPsynthase
NO3–
SO42–
Electron Transport Chain
•A chain of special redox carriers that receives reduced carriers (NADH, FADH2) generated by glycolysis and the Krebs cycle
- passes them in a sequential and orderly fashion from one to the next
- highly energetic- allows the transport of hydrogen ions outside of the
membrane- in the final step of the process, oxygen accepts electrons and
hydrogen, forming water- Electron transport carriers and enzymes are embedded in the
cell membrane in prokaryotes and on the inner mitochondrial membrane in eukaryotes
Electron Transport Chain
•Released energy from electron carriers in the electron transport chain is channeled through ATP synthase
•Oxidative phosphorylation: the coupling of ATP synthesis to electron transport
- each NADH that enters the electron transport chain can give rise to 3 ATPs
- Electrons from FADH2 enter the electron transport chain at a later point and have less energy to release, so only 2 ATPs result
Electron Transport Chain
- electrons from the chain and H+ from solution react with oxygen to form water
2H+ + 2e‐ + ½ O2 H20
Final Electron Acceptor: Aerobic Respiration
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•Most eukaryotes have a fully functioning cytochrome system
•Bacteria exhibit wide‐ranging variations in this system- some lack one or more redox steps
- several have alternative electron transport schemes
- lack of cytochrome c oxidase is useful in differentiating among certain genera of bacteria (we will do this with MM)
Final Electron Acceptor: Aerobic Respiration
• Utilizes oxygen‐containing ions, rather than free oxygen, as the final electron acceptor
Ex. Nitrate reductase
NO3‐ + NADH NO2
‐ + H2O + NAD+
•Nitrate reductase catalyzes the removal of oxygen from nitrate, leaving nitrite and water as products
Final Electron Acceptor: Anaerobic Respiration
Table 7.5
Pyruvic acid from glycolysis can itself become the electronacceptor.
Pyruvic acid can also be enzymatically altered and then serve asthe electron acceptor.
The NADs are recycled to reenter glycolysis.
The organic molecules that became reduced in their role aselectron acceptors are extremely varied, and often yield usefulproducts such as ethyl alcohol, lactic acid, propionic acid,butanol, and others.
C C
O
H
H
H
H
CC C
H
H
H
H
O
C C
H
H
H
H
H
C C C
Lactic acid
OH
OH
NAD+
Ethyl alcohol
OH
Acetaldehyde
CO2
Pyruvic acid
Remember: Thishappens twice for
each glucosemolecule that
enters glycolysis.
Fermentation
NAD H NAD H
Fermentation- the incomplete oxidation of glucose or other carbohydrates in the absence of oxygen
- uses organic compounds as the terminal electron acceptors
- Yields 2 ATPs per molecule of glucose
- Used by organisms that do not have an electron transport chain
- Other organisms repress the production of electron transport chain proteins when oxygen is lacking in their environment to revert to fermentation
- Many bacteria grow as fast as they would in the presence of oxygen due to an increase in the rate of glycolysis
•Permits independence from molecular oxygen- allows colonization of anaerobic environments- enables adaptation to variations in oxygen availability- provides a means for growth when oxygen levels are too low for
aerobic respiration
Fermentation
•Bacteria and ruminant cattle- digest cellulose through fermentation
- hydrolyze cellulose to glucose
- ferment glucose to organic acids which are absorbed as the bovine’s principal energy source
Fermentation Final Electron Acceptor: Fermentation
- Uses organic compounds as the terminal electron acceptors
• Products:• Alcoholic beverages:
ethanol and CO2
• Solvents: acetone, butanol
• Organic acids: lactic acid, acetic acid
• Vitamins, antibiotics, and hormones
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Differential and Selective Media: Mannitol Salt Agar
S. epidermidis S. aureus
• Physiological reactions: indirect evidence of enzymes present in a
species. If bacteria tests + for oxidase that means it uses cytochrome c oxidase in ETC. If it is negative it means it uses another
enzyme for ETC or it does not go through ETC.
Phenotypic Methods: Biochemical Testing
Energy present in the electron acceptor can be captured to phosphorylate ADP to store energy in ATP
Oxidation and Reduction