• At this point, pyruvate can either be used:
– in fermentation to form fermentation products (alcohols, acids, gases) OR
– in respiration (TCA cycle) to be fully oxidized to carbon dioxide
Fermentation vs. Respiration
• Fermentation
– from 1 glucose molecule a net of 2 ATP are produced
– Partial oxidation of glucose
– reduction of pyruvate
• fermentation products
• NAD+
• Respiration
– from 1 glucose molecule up to 38 ATP are produced
– complete oxidation of glucose to CO2
– oxidation of pyruvate by citric acid cycle
– uses electron transport chain and ATPase
Fermentation
• During fermentation, glycolysis occurs followed by fermentation reactions. (no TCA, no ETC)
• During glycolysis, electrons from glucose are passed to 2 NAD+ creating 2 NADH
• In fermentation, electrons from NADH are passed to pyruvate, regenerating NAD+ so glycolysis can continue
• In this process (glycolysis followed by fermentation reactions), net gain of 2 ATP are made by substrate-level phosphorylation
Types of fermentation
1. Homolactic fermentation- once electrons from NADH are passed to pyruvate, pyruvate is reduced to lactic acid
• Ex. Lactobacillus ferments lactose sugar in milk to produce lactic acid, this gives us yogurt
2. Alcoholic fermentation- once electrons are passed from NADH to pyruvate, pyruvate is reduced to alcohol and CO2
• Ex. Yeast ferment the sugar in malted grains to produce alcohol & CO2 (beer, wine, bread)
• In both types of fermentation, the electron donor is glucose & electron acceptor is pyruvate, which is made from glucose.
• Fermentation- energy generating process where one organic compound (ex.glucose) serves as the electron donor and a product of that compound serves as the electron acceptor (ex.pyruvate).
Respiration
• Some chemoheterotrophs can use oxygen or other compounds from the environment to accept the electrons from NADH.
• If electron acceptor is oxygen, process is called aerobic respiration
• If electron acceptor is other compound from environment, process is called anaerobic respiration
– final electron acceptor may be an inorganic compound such as nitrate, ferric iron, sulfate or carbonate.
• In fermentation, the final electron acceptor is made by the cell from the electron donor
– (ex. Pyruvate made from glucose)
• In respiration, the final electron acceptor is a compound found in the environment
– (ex. Oxygen, nitrate)
The tricarboxylic acid (TCA) cycle
• aka- Kreb’s cycle, citric acid cycle
• cyclic pathway used to fully oxidize organic materials like acetyl CoA
• End products:
– ATP (small amount)
– NADH and FADH2 (important, used in ElectronTransportChain)
– CO2 (waste product)
– Precursor metabolites
• Occurs during respiration
• _Oxidizes_ completely to CO2
• Only possible with inorganic electron acceptor
• Respiration produces more energy than fermentation
• _Pyruvate dehydrogenase_ complex (PDC)
– Converts pyruvate to acetyl-CoA by removing CO2
• Substrate for TCA is acetyl CoA (made by removing CO2 from pyruvate), NOT pyruvate.
• Precursor metabolites made in TCA cycle:
– Alpha-ketoglutarate & oxaloacetate (used to make amino acids & nucleotides)
Reactions of the TCA cycle
• Acetyl CoA + oxaloacetate= citrate (6 C compound)
• During this cycle, acetyl CoA is oxidized to CO2 & original oxaloacetate is regenerated
• _CO2_ released
• _NADH and FADH_ are generated
• precursor metabolites are made to be used for biosynthesis
See Fig 13.27
Aromatic Catabolism
• Bacteria can degrade many compounds
– Pseudomonas, Rhodococcus
• _Aromatic_ compounds converted to pyruvate which enters TCA cycle
– Allows growth in wide range of environments
– Used for bioremediation (using microorganisms to detoxify)
· Cleaning up oil spills
· Cleaning industrial sites
· Degrading toxic compounds
Aerobic respiration
• In aerobic respiration, the pair of electrons from NADH is passed through a series of intermediates to oxygen
• NADH is oxidized back to NAD+, oxygen is reduced to water
• Oxygen is the terminal electron acceptor
• There are several types of intermediates that pass electrons from NADH to oxygen:
– flavoproteins
– cytochromes
– quinones
– iron-sulfur proteins
• These components make up an electron transport chain (ETC)
• In prokaryotes, the ETC is in the cytoplasmic membrane
• In eukaryotes, the ETC is in the mitochondria
• Electron transport chain- series of membrane associated electron carriers that carry electrons from the primary electron donor to the terminal electron acceptor
• During electron transfer, electron transport systems conserve some of the released energy for the synthesis of ATP
– ETC proteins are called oxoreductases because they oxidize one substrate then reduce another
Electron transport chain
• Different organisms have different ETC
• Some have more than one for different growth conditions
• Electrons are passed from one compound to the next
• At each step, some energy from the electrons is used to push hydrogen ions across the cytoplasmic membrane into the periplasm or to outside of CM
Hydrogen ion gradient/proton motive force/chemiosmosis
• This creates a hydrogen ion gradient across the CM
• Outside of CM is now more acidic and more + than the cytoplasm
• Some organisms such as extreme halophiles use sodium motive force (Na+ ion gradient)
• This gradient is a source of potential energy
student.ccbcmd.edu/biotutorials/cellresp/images/chemios.gif ATPase
• ATPase- enzyme which adds a phosphate group to ADP to make ATP
• ATPase does this by allowing protons to cross back into the cytoplasm
• Energy is released by reducing the hydrogen ion gradient, this energy is used to make the high energy bond of ATP.
• Oxidative phosphorylation- process of using the hydrogen ion gradient to make ATP
Substrate-level vs. Oxidative Phosphorylation
• Substrate-level phosphorylation
– ATP is made by transferring a high energy phosphate group to ADP
• Oxidative phosphorylation
– Energy from a hydrogen ion gradient is used to make ATP
Respiration and Fermentation summary
• Fermentation- glucose to pyruvate to fermentation products--electrons from NADH used to reduce pyruvate. Net 2 ATP/glucose.
• Respiration- glucose to pyruvate then pyruvate is oxidized to CO2 in TCA cycle-electrons from NADH are transferred to oxygen through ETC. Up to 38 ATP/glucose.
Anaerobic respiration
• Same process as aerobic respiration
• Electrons from NADH are passed down an ETC, losing energy at each step
• Energy is used to move protons across cytoplasmic membrane to periplasm
• Energy from hydrogen ion gradient is used by ATPase to make ATP (oxidative phosphorylation)
• Difference- final electron acceptor is some compound OTHER THAN oxygen
Gluconeogenesis- production of glucose from nonsugar precursors.
• Some cells can make glucose from _amino acids, glycerol and fatty acids_ using this pathway.
• _Glycolysis in reverse_. Uses most of the same enzymes as glycoysis. There are 4 unique enzymes used in this pathway.
• Requires large amounts of energy
Biosynthesis
Anabolism
• _11 precursor metabolites_ in E. coli
– 6 from glycolysis, 3 from TCA, 2 from pentose phosphate
• Anabolic pathways- convert precursors (made in glycolysis, TCA, etc) into building blocks (monomers) such as amino acids & nucleotides
• Monomers are polymerized to form macromolecules (proteins & nucleic acids) & structures (LPS).
Monomer
Polymer
Sugar
polysaccharide
Nucleotide
nucleic acid
Amino acid
protein
Fatty acid
lipid
For organisms growing in an environment without building blocks, they must be biosynthesized from simpler components, a process called anabolism.
Metabolic Diversity
The Phototrophic Way of Life
Phototrophs- use
2 types of photosynthesis:
• Anoxygenic- no oxygen
– Most photosynthetic bacteria are anoxygenic phototrophs
• Oxygenic- water is split to produce
– ___________________________________________________________ are oxygenic phototrophs
Pigments of photosynthesis:
• chlorophylls in oxygenic phototrophs
• bacteriochlorophylls in anoxogenic phototrophs
• located in _chloroplasts_ where the light reactions of photosynthesis are carried out
• Photosynthetic eukaryotes have _chloroplasts_ that contain the photosynthetic membranes known as _thylakoids_
• Since photosynthetic prokaryotes do not have chloroplasts, the photosynthetic membranes are:
• the _cytoplasmic membrane_ in many bacteria
• _chlorosomes_ in green bacteria or
• _thylakoid membranes_ in cyanobacteria
• The ultimate in low-light efficiency is found in the _chlorosome_ of green sulfur bacteria and Chloroflexus.
• Can grow at lowest light intensities of all known prokaryotes
• _Antenna chlorophyll_ molecules harvest light energy and transfer it to _reaction center chlorophylls_ where the conversion of light energy to ATP occurs.
• Different pigments absorb different wavelengths of light
• There are several different chlorophylls and bacteriochlorophylls, each with a unique absorption spectra
• Different species have different pigments making it easy for two organisms to coexist in a habitat without competing for light energy.
• carotenoids and phycobilins- _accessory pigments_ that absorb light and transfer the energy to reaction center chlorophylls.
• Chlorophylls can only absorb certain wavelengths of light. Accessory pigments allow organisms to capture additional wavelengths of light
• Carotenoids also play an important _photoprotective_ role in preventing _photooxidative damage_ (due to toxic forms of oxygen) to cells.
• In photosynthesis, a series of electron transport reactions in the photosynthetic reaction center of phototrophs results in the formation of a proton motive force and the synthesis of ATP.
Anoxygenic Photosynthesis
• Photosynthesis that does not produce oxygen
• Anoxygenic phototrophs include members of the following bacterial phyla:
· Proteobacteria (purple sulfur & purple nonsulfur)
· Chloroflexus (green nonsulfur)
· Chlorobium (green sulfur)
· Heliobacteria (gram +)
· Rhodobacter species are used for studying anoxygenic photosynthesis
• They use photosynthesis in the light, respiration in the dark, & grow in the presence or absence of O2.
Oxygenic Photosynthesis
• Algae and cyanobacteria use electrons from H2O to reduce NADP for CO2 fixation, producing O2 as a byproduct.
• Two separate light reactions in oxygenic photosynthesis, photosystems I & II.
• Photosystem I resembles the system in anoxygenic photosynthesis.
• Photosystem II splits H2O to yield O2
Autotrophic Fixation: The Calvin Cycle
• Autotrophs- use CO2 as sole carbon source
• Convert CO2 into organic carbon compounds (CO2 fixation)
• Many use the Calvin-Benson cycle
· Calvin cycle- _Rubisco enzyme catalyzes condensation of CO2 + ribulose 1,5 bisphosphate_ to make 3-phosphoglycerate then glyceraldehyde 3-phosphate then fructose-6-phosphate
• requires large amount of ATP
• F-6-P goes into glycolysis
• Glycolysis & TCA occur just as mentioned previously starting with F-6-P.
• Several autotrophic prokaryotes that use the Calvin cycle for CO2 fixation produce cell inclusions called carboxysomes that store molecules of RubisCO.
• The Calvin cycle is an energy-demanding process in which CO2 is converted into sugar.
• Hexose sugars can be used by the cell or put into storage polymers like glycogen, starch, or poly-β-hydroxyalkanoates.
Autotrophic Fixation: Reverse TCA Cycle and the Hydroxypropionate Cycle
• Green sulfur bacteria use the reverse TCA cycle for CO2 fixation
• Green nonsulfur bacteria such as Chloroflexus use the hydroxypropionate pathway
• Chloroflexus is the oldest anoxygencic phototrophic Bacteria
• Possibly the first attempt at autotrophy in anoxygenic phototrophs
Chemolithotrophy: Energy from the Oxidation of Inorganic Electron Donors
• Chemolithotrophs- _oxidize inorganic_ chemicals as their sole source of energy
• Most are also autotrophs
• Some are _mixotrophic_- although they are able to obtain _energy_ from the oxidation of an _inorganic_ compound, they require an _organic_ compound as a _carbon_ source.
• Use inorganic compounds such as hydrogen sulfide, hydrogen gas, ferrous iron, and ammonia as electron donors
• Most of the time _oxygen_ is the terminal electron acceptor; so aerobic respiration takes place, it just starts with an inorganic instead of organic molecule.
Examples of chemolithotrophy:
1. Hydrogen Oxidation
Hydrogen bacteria:
• _Oxidize H2_, thereby generating a proton motive force and ATP synthesis.
• These chemolithotrophs are also autotrophs and fix CO2 via the Calvin cycle
• Generate energy by oxidizing hydrogen gas
• Electrons from _hydrogen_ are passed down an ETC to oxygen (aerobic respiration)
• Hydrogen ion gradient in formed
• ATPase uses potential energy from this gradient to make ATP
• Example- Ralstonia
• If organic compounds are present, most hydrogen bacteria will grow as _chemoorganotrophs_, but if organic compounds are absent they grow chemolithotrophically and fix CO2 by the Calvin cycle.
2. Oxidation of Reduced Sulfur Compounds
Sulfur bacteria:
• Oxidize reduced sulfur compounds such as H2S and S0 to sulfate (SO42-)
• Sulfate is exported to the environment where it reacts with hydrogen ions to form sulfuric acid
• When sulfur is oxidized, electrons are donated to ETC where electron acceptor is usually oxygen (aerobic)
• Causes hydrogen ion gradient which ATPase uses to generate ATP
• These chemolithotrophs are also autotrophs & use the Calvin cycle to fix CO2.
• Most are aerobic
• Example- Beggiatoa
3. Iron Oxidation
Iron oxidizing bacteria:
• Chemolithotrophs that generate energy by oxidizing ferrous iron (Fe2+) to ferric iron (Fe3+)
• Most are obligately acidophilic.
4. Nitrification
Nitrifying bacteria:
• NH3-->NO3-
• oxidize inorganic nitrogen aerobically by process called nitrification
• Ammonia (NH3) and nitrite (NO2-) are the most common inorganic nitrogen compounds used as electron _donors_
• Nitrosifyers oxidize ammonia to nitrite which is then oxidized by nitrite oxidizers to nitrate.
• Most nitrifying bacteria are also _heterotrophs_ - able to use glucose or other organic carbon source.
Metabolic Diversity
Anaerobic Respiration
• Anaerobic respiration
– uses molecules other than oxygen as final electron acceptor
• Obligate anaerobes solely use anaerobic respiration
• Some facultative anaerobes like denitrifying bacteria use aerobic respiration in the presence of oxygen but when oxygen is depleted switch to anaerobic respiration.
1. Nitrate Reduction and Denitrification
Denitrifying bacteria- Ex. pseudomonas
· Reduce nitrate (NO3-) to gases such as nitrogen (N2), nitrous oxide (N2O) and nitric oxide (NO)- (escape from environment
· nitrate is the electron acceptor
· ETC process is same as aerobic respiration
· Difference- electrons are used to reduce nitrate instead of oxygen
2. Sulfate Reduction
• Sulfate-reducing bacteria use sulfate as an electron acceptor (anaerobic respiration), converting the sulfate to hydrogen sulfide
How would the above ETC diagram be different for sulfate reducers?
3. Methanogenesis
• Biological production of methane (CH4) from CO2 plus H2 or from methylated compounds
• Methanogens- strictly _anaerobic Archaea_ capable of methanogenesis
• Energy is produced using proton and sodium ion gradients
• CO2 is final electron acceptor
Besides inorganic nitrogen and sulfur compounds or CO2, a variety of other substances, both organic and inorganic, can function as electron acceptors for anaerobic respiration
Metabolic Diversity
Nitrogen Fixation
· Nitrogen fixation is the reduction of atmospheric N2 to ammonia (NH3).
• Most organisms must use oxidized nitrogen compounds (NO3-, NH3) from environment as nitrogen source
• Some can use nitrogen gas (nitrogen fixation)--convert it into NH3 which is used to make organic compounds
• 40 ATPs consumed for each N2 fixed
• Occurs in some prokaryotes, not in eukaryotes
• Nitrogen fixation requires an enzyme complex called nitrogenase
• Nitrogenase is inhibited by oxygen
• Many organisms only fix N when growing anaerobically
• Aerobic nitrogen-fixing bacteria prevent the oxygen needed for respiration from interacting with nitrogenase by:
– protective proteins to stabilize nitrogenase
– rapid removal of O2 by respiration
– producing O2 retarding slime layers
– compartmentalization of nitrogenase in special cells (some species of cyanobacteria have heterocysts where nitrogen fixation occurs)
– Others (Rhizobium) are intracellular symbionts within plants (legumes)
– Rhizobium provides plants with N, plant protects bacteria and keeps nitrogenase from being destroyed by oxygen