Metabolism = breaking molecules down and building up new ones

Important processes in metabolism Discuss processes in order in which they (might have) evolved

1. Anaerobic breakdown of organic molecules = fermentation. Fits with primordial soup argument (first organisms heterotrophic). 2. Respiration electron transport chains (still heterotrophs but much more efficient). 3. Chemosynthesis (autotrophs can carry out carbon fixation. No longer limited by the soup). 4. Photosynthesis (autotrophs huge amounts of energy for free! Major increase in biomass).

Glycolysis breakdown of sugar

Essentials worth remembering 1 glucose (6C) 2 pyruvate (3C) Generates 2 ATP and 2 NADH

Essentials In anaerobic bacteria pyruvate is broken down to waste products (e.g. lactate). NAD+ is regenerated (a cycle)

also occurs in muscles

Other examples of fermentation processes:

CO2 Pyruvate CO2 + ethanol; These occur in yeast Glucose is only partly oxidized by these reactions. Relatively inefficient. Pyruvate CO2 + acetic acid

In aerobic organisms, pyruvate feeds into the Citric Acid Cycle (Krebs cycle)

Acetyl CoA

Essentials This produces NADH and FADH2. These are electron donors (reducing agents) for the electron transport chain. All the C from the glucose is now oxidized to CO2. Many other biosynthetic pathways branch off from glycolysis and citric acid cycle.

Important processes in metabolism Discuss processes in order in which they (might have) evolved

1. Anaerobic breakdown of organic molecules = fermentation. Fits with primordial soup argument (first organisms heterotrophic). Relatively simple. 2. Respiration electron transport chains (still heterotrophs but much more efficient). Really clever, but complicated.

3. Chemosynthesis (autotrophs can carry out carbon fixation. No longer limited by the soup).
4. Photosynthesis (autotrophs huge amounts of energy for free! Major increase in biomass).

Oxidation-Reduction again Nicotinamide adenine dinuclotide NAD+

FAD
Flavin adenine dinucleotide

oxidizing agent
(electron acceptor)

FADH2

NADH reducing agent FADH2 FAD + 2H+ + 2e(electron donor) NADH NAD+ + H+ + 2eNow we are going to make use of those electron donors we just made two slides back. Hang onto your hats!

H+

H+

H+

2eNADH H+ NAD+ NADH dehydrogenase complex Essentials Aerobic respiration (in aerobic bacteria or in mitochondria in eukaryotes) heme group in cytochrome c cytochrome b-c1 complex

ubiquinone

H+

cytochrome c

H2O H+ 2H+ + O2

cytochrome oxidase complex

High energy electron donor eventually donates electrons to O2

Electron goes downhill in G Proton gradient is generated.

ATP synthetase complex

proton channel

Electron transport chain + ATP synthesis = oxidative phosphorylation chemiosmotic process For each molecule of glucose about 30 ATPs generated by ox. phos. but only 2 from glycolysis. Much more energy from the same food! ADP + Pi ATP protons moving downhill provide energy for uphill synthesis of ATP

Other respiratory chains In each case organic molecules are oxidized. The terminal electron acceptor is reduced. The energy released is used to generate a proton gradient that is used for ATP synthesis. In aerobic respiration O2 is the electron acceptor. In anaerobic respiration another molecule is the electron acceptor.
Type of metabolism Aerobic respiration Denitrification Electron acceptor O2 NO3Products H2O NO2-. NO2-,, N2O or N2 H2S H2S Organisms Many aerobic bacteria and archaea. Eukaryotes (mitochondria) Many bacteria can do this facultatively (eg. E. coli, B. subtilis). Paracoccus denitrificans (B) Desulfovibrio desulfuricans (B) Archaeoglobus fulgidus (A#) Delsulfuromonas acetoxidans (B) Pyrococcus, Desulfurococcus (A) Sulfolobus, Thermoproteus (A#)

Sulphate reduction Elemental sulphur metabolism

SO42S (red. with H2)

Iron reduction

Fe3+

Fe2+

Thermus (B)

A Archaea; B Bacteria; # can also be chemoautotrophic

Evolution of respiratory chains

Early organisms probably used fermentation only (anaerobic).

Fermentation usually leads to excretion of acids (lactic, formic, acetic....). Proton pump would be favoured to keep the acid out. H+

ATP synthase works both ways. May have originated as an ATP driven proton pump.
H+ ATP ADP + Pi

Electron transport chain enabled H+ to be pumped without using ATP.

e-

H+ If electron transport chain pumps became more efficient than necessary, the proton gradient could be used to drive ATP synthase to make ATP.

eH+ ADP + Pi ATP

Important processes in metabolism Discuss processes in order in which they (might have) evolved 1. Anaerobic breakdown of organic molecules = fermentation. Fits with primordial soup argument (first organisms heterotrophic). Relatively simple. Maybe these kind of reactions were catalyzed by ribozymes in the RNA world. NADH, FADH2, CoA all involve nucleotides (clue?). 2. Respiration electron transport chains (still heterotrophs but much more efficient). Really clever, but complicated. Each complex in the respiratory chain involves many proteins. No RNAs known to do this. probably this comes after RNA world but before LUCA Now we can efficiently generate energy from food, but we are running out of food... 3. Chemosynthesis (autotrophs can carry out carbon fixation. No longer limited by the soup). 4. Photosynthesis (autotrophs huge amounts of energy for free! Major increase in biomass).

Chemoautotrophy (Chemolithotrophy)

An inorganic reducing agent feeds into an electron transport chain. Generates a proton gradient (more ATP synthesis) and an organic reducing agent (like NAD(P)H), which reduces CO2 to organic molecules. Several different carbon fixation cycles are known opposite of citric acid cycle.
Type of metabolism
Hydrogen oxidation Nitrification (from nitrite or ammonia) Sulphur oxidation (from thiosulphate, sulphur or hydrogen sulphide) Iron oxidation Methylotrophy Methanogenesis Elemental sulphur metabolism Sulphate reduction

Energy producing reaction

H2 + O2 H2O NO2- + O2 NO3NH4+ + 1 O2 NO2- + H2O + 2H+ S2O32- + 2O2 + H2O 2SO42- + 2H+ S + 1 O2 + H2O SO42- + 2H+ 2H2S + O2 2S + 2H2O 2Fe2+ + 2H+ + O2 2Fe3+ + H2O CH4 or CH3OH or CO CO2 4H2 + CO2 CH4 + 2H2O H2 + S H2S H2 + SO42- (or SO32- or S2O32-) H2S

Organisms
Alcaligenes, Hydrogenobacter (B) Nitrobacter(B) Nitrosomonas (B) Sulfolobus (A) Thiobacillus (B) Thiobacillus (B) Thiobacillus (B) Methylomonas (B) Methanococcus (A) Thermoproteus (A) Archaeoglobus (A)

Essentials

Many possible energy sources from redox reactions.

Can go both ways - 2 examples: can oxidize S to SO42- in aerobic conditions or reduce S to H2S in presence of H2 gas but absence of O2 ---- both have G < 0 in the right conditions.

methylotrophy (aerobic) v. methanogenesis (anaerobic)

Sometimes the same organism goes both ways:

e.g. Sulfolobus can be an anaerobic heterotroph with sulphur reduction, or an autotrophic aerobic sulphur oxidizer clever cloggs!

Redox reactions in previous table have G < 0. They look simple, but remember they dont just happen in one step as an inorganic reaction. These reactions are coupled to electron transport chains and proton gradients....

Important processes in metabolism Discuss processes in order in which they (might have) evolved 1. Anaerobic breakdown of organic molecules = fermentation. Fits with primordial soup argument (first organisms heterotrophic). Relatively simple. Maybe occurred in the RNA world.

2. Respiration electron transport chains (still heterotrophs but much more efficient). Really clever, but complicated. Each complex in the respiratory chain involves many proteins. No RNAs known to do this. probably this comes after RNA world but before LUCA
3. Chemosynthesis (autotrophs can carry out carbon fixation. No longer limited by the soup). Many possible sources of chemical energy. Some of these types of metabolism are found in both archaea and bacteria, i.e. before LUCA. 4. Photosynthesis (autotrophs huge amounts of energy for free! Major increase in biomass). Only in bacteria, i.e. after LUCA requires light-harvesting protein complexes (photosystems)

Complementary processes of photosynthesis and respiration Carbon fixation into sugars reduction of CO2 Oxidation of sugars into CO2

(Some forms of photosynthesis do not produce oxygen)

(In anaerobic organisms sugars are oxidized incompletely via fermentation. O2 not required.)

Two types of chlorophyll absorb visible light at slightly different wavelengths. Chlorophyll contained in the photosystem I and II protein complexes

high energy electron enters the transport chain

light excites an electron delocalized electrons in ring structure

low energy electron replaces it

Photosynthesis: a light-driven electron transport chain Thylakoid membrane of chloroplasts (or outer membrane of photosynthetic bacteria) light H2O light

2H+ + O2

H+

2e-

plastoquinone H+ Photosystem II

plastocyanin

ferredoxin NADP+

NADPH

cytochrome b6-f complex Generates proton gradient that can be used by ATP synthase

Photosystem I

FerredoxinNADP reductase

NADPH is a reducing agent that can reduce CO2 to organic molecules

The dark reactions of photosynthesis.

Carbon fixation cycle (Calvin cycle).

CO2 is reduced to sugars. Requires energy and reducing power.

Types of photosynthesis

5 groups of bacteria perform photosynthesis.

In oxygenic photosynthesis H2O is the electron donor and O2 is produced. In anoxygenic photosynthesis H2S is the electron donor and O2 is not produced.
Type of photosynthesis
Anoxygenic Anoxygenic Anoxygenic

Photo-system
PS I PS I PS II

Organisms
Green sulphur bacteria - Chlorobium Heliobacteria Purple sulphur bacteria (Chromatiales Gamma proteobacteria) Purple non-sulphur bacteria (Rhodospirillum Alpha proteobacteria). Use H2 not H2S Green filamentous bacteria - Chloroflexus Cyanobacteria and Chloroplasts (in Eukaryotes)

Anoxygenic Oxygenic

PS II PS I and PS II

Evolution of photosynthesis (see Olsen and Blankenship, 2004)

PS I Chlorobium and Heliobacteria

divergence in separate lineages

ancestral PS

fusion

PS I & II Cyanobacteria

endosymbiosis: chloroplasts

PS II Chloroflexus and Purple bacteria

PSs contain different types of chlorophyll. Genes for pigment synthesis may not follow same tree as genes for the components of the PSs. Evidence for horizontal transfer. Archaea do not have these photosystems. They evolved after the LUCA. However: Halobacteria (which are salt-loving extremophile archaea) have an independent light harvesting protein called bacteriorhodpsin in their purple membrane. Contains retinal chromophore. Different to chlorophyll.

Archaea

Eukaryotes
chloroplasts

Bacteria

Plausible summary of Everything

Methanogenesis/ Bacteriorhodopsin only in Archaea

Oxygenic Photosynthesis Anoxygenic Photosynthesis

mitochondria origin of eukaryotic nucleus ?

LUCA

Genes for sulphate reduction, nitrate reduction, sulphur oxidation, oxygen respiration all present in A and B

Modern organisms: DNA + RNA + proteins Genetic code: RNA + proteins RNA world Metabolism first ? Origin of life

Chemosynthesis
Electron transport chains

Simple heterotrophic metabolism / fermentation

Alternative viewpoint # 1 Early evolution of photosynthesis Mauzerall argues that only photosynthesis could supply sufficient energy for life. Light absorbing pigments must have existed very early. These would have initiated redox reactions. But these would be independent of todays membrane bound electron transport chains. ?? But some proteins in the respiratory and photosynthetic chains are related. Suggests that (current form of) photosythesis was later. Alternative viewpoint # 2 Chemoautotrophic origin Wchtershuser argues that an autotrophic metabolism based on pyrite was first. FeS + H2S FeS2 + H2 ?? This may be a plausible energy source but (current forms of) autotrophs use complex electron transport pathways. If this existed, evidence of it is lost. ?? The first organisms must have been made of something! Presumably organic molecules .... This brings us back to the primordial soup.... Alternative viewpoint # 3 Clay mineral origin Cairns-Smith argues that organic molecules were not important originally. Clay minerals stored information. Genetic takeover occurred (e.g. to RNA).

Extremophiles
What counts as extreme? Depends on our viewpoint. What limits organisms? Challenges in different environments. How to overcome them?

What can they tell us about possibility of life elsewhere?

Congress pool. Yellowstone. pH3 80oC

Sulfolobus acidocaldarius

Pictures from Rothschild & Mancinelli (2001) See also Lunine Chap 10 Chapters by Rothschild and Stetter in OI book.

Temperature

>80 Hyperthermophiles

60-80 Thermophiles
15-60 Mesophiles <15 Psychrophiles Eukaryotes more limited at high temp than bacteria and archaea Low temp organisms from all domains Growth rate measurements distinguish tolerant organisms from true philes

Challenges of high T:

stability of molecular structures, membranes, and molecules themselves

Examples of molecular adaptation to high T: GC content in rRNA correlated with growth temp. (Galtier & Lobry, 1997) Higher helix melting temp In proteins Gunfolding found to be large in thermozymes Tunfolding is higher More hydrogen bonds with water. More salt bridges between + and charged residues. More disulphide bonds between cysteines. Folded structures more rigid, fewer cavities.

10

60

110

But overall genomic GC content does not correlate with T. DNA must be stable anyway...

Psychrophiles challenges of low temps Membrane becomes too rigid need to change lipid structure Slows down reaction rates Liquid water usually required for reactions Ice crystals expand relative to water can tear cells apart. Antifreeze proteins found in fish that live at < 0 Small helical proteins can bind to the surface of small ice crystals and prevent them growing.

Sea-ice diatoms (unicellular photosynthetic eukaryotes)

Salinity Halophiles Salt conc in ocean is 3.5%, but this is too high for us. Some organisms are adapted to concs up to 35% in salt lakes

Water will diffuse out of the cell by osmosis. Causes dessication. Many halophiles use Compatible solutes - small organic molecules that do not interfere with metabolism when accumulated to high conc. Extreme halophiles use salt-in-cytoplasm K+ are selectively allowed into cell to balance the osmotic pressure. Enzymes have to adjust to working in this situation.

Halobacteria in a salt lake (Archaea with photosynthetic purple membrane)