Physiology and Diversity of Prokaryotes WS 2009/2010 ( PHOTOTROPHS. Martin Könneke. Lithotrophic Processes

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1 Physiology and Diversity of Prokaryotes WS 2009/2010 ( PHOTOTROPHS Martin Könneke Lithotrophic Processes Elektronendonor Oxidized product Process/ organism H 2 H + (H 2 O) Knallgas reaction/ Ralstonia NH + 4 NO - 3 Nitrification (2 types) NH + 4 NO - 2 Ammonia oxidizer (Nitroso-) NO - 2 NO - 3 Nitrite oxidizer (Nitro-) CH 4 CO 2 Methane oxidizer (Methylo-) H 2 S, S SO 2-4 Sulfur oxidizer/thiobacillus, Beggiatoa Fe 2+ Fe 3+ Iron oxidation/thiobacillus H 2 O O 2 Photosynthesis

2 Lithotrophic processes are essential for the reoxidation of reduced electron acceptors! All chemolithotrophes are prokaryotes! Almost all known lithotrophes are autotroph! Energieform Elektronendonor Kohlenstoffquelle Chemo- Organo- heterotroph Photo- Litho- autotroph

3 CO 2 fixation pathways At present, 6 different pathways are known, just a single one within the Eukarya Differ with regard to energy requirement, end products and oxygene sensitivity Calvin Cycle Reductive (reverse) citric acid cycle Reductive acetyl-coa pathway 3-Hydroxypropionate cycle ( 2 variations)

4 CO 2 fixation: Calvin Cyclus Most widespead carbon fixation pathway (RubisCO most abundant enzyme on Earth) Occurs in chloroplast, cyanobacteria, and most chemolithoautotrophic bacteria Some bacteria contain speciallized compartements, carboxysome, with high RubisCO concentration Reduces CO 2 even at high oxygen concentrations Can also funtion as oxygenase CO 2 fixation via the Calvin Cyclus (Calvin-Bassham-Benson-cylcle) Key enzyme: RubisCO Ribulosebisphosphat-Carboxylase/Oxygenase Used by all plants, cyanobacteria, and most of the aerobic chemolithoautotrophic bacteria Reduction of CO 2 to the oxidation state of sugar: CO ATP + 4[H]! <CH 2 O> + H 2 O + 3 ADP + 3 P i - IV 0

5 CO 2 Fixierung: Calvin Cyclus RubisCO CO 2 Fixierung: Calvin Cyclus 15 C 3 C Phosphoribulokinase 3 C 18 C 15 C RubisCO

6 CO 2 fixation: Calvin Cyclus Key enzyme: RubisCO Ribulosebisphosphat-Carboxylase/Oxygenase Requirements for the synthesis of 3-phosphate glycerine aldehyde 3 CO ATP + 6 NADPH carbon energy reducing power Energy expensive pathway! Reductive (reverse) citric acid cycle Represents the reversion of the citric acid cycle Replacement of 3 enzymes: 1) ATP-citrate lyase instead of the citrate synthase 2)!-ketoglutarate-synthase instead of "- ketoglutaratedehydrogenase 3) Fumarate synthase instead of succinate dehydrogenase Final product of the cycle is acetyl-coa, that is further carboxylized to pyruvate. A third step is the ATP dependent conversion to triose-phosphate.

7 Reductive citric acid pathway Reductive citric acid pathway

8 Reductive (reverse) citric acid cycle Present in: Phototrophic green sulfur bacteria (e.g. Chlorobium limicola) Sulfate reducers (Desulfobacter hydrogenophilus) Knallgas bacteria (Hydrogenobacter thermophilus) Thermophilic, sulfur-reducing archaea (Thermoproteus neutrophilus) The acetyl-coa pathway - In contrast to other carbon fixation pathways, not a cycle - two linear reaction series resulting in A) a methyl- and B) a carbonyl group - key enzyme: CO-DH (Carbon monooxide dehydrogenase) CO 2 + H 2! CO + H 2 O - The CO 2 reduction must be considered as bifunctional pathway: A) energy metabolism B) C-fixation for biosynthesis

9 Reductive acetyl-coa pathway

10 3-Hydroxypropionate cycle Reduction of bicarbonate to gyoxylate Bicarbonate fixing enzymes are: acetyl-coa carboxylase and propionyl-coa carboxylase 3-hydroxypropionyl-CoA as characteristic intermediate Recycling of the primary carbonate acceptor acetyl-coa 3-Hydroxypropionate cycle

11 3-Hydroxypropionate cycle 3-Hydroxypropionate cycle At present only found in the green nonsulfur phototrophic members of the genus Chloroflexus and in thermophilic Crenarchaeota (Metallosphaera) Suggested to be oldest pathway of autotrophy in anoxygenic phototrophes

12 Acetyl-CoA Acetyl-CoA HCO - 3 Acetyl-CoA carboxylase (Nmar_0272, 0273, 0274) Malonyl-CoA Malonyl-CoA reductase Malonate semialdehyde reductase (unknown) Succinyl-CoA Methylmalonyl-CoA 3-Hydroxybutyryl-CoA Crotonyl-CoA hydratase (Nmar_1308) Crotonyl-CoA 3-Hydroxypropionate 3-Hydroxypropionyl-CoA synthetase 4-Hydroxybutyrate 3-Hydroxypropionyl-CoA dehydratase Acryloyl-CoA reductase (unknown) Propionyl-CoA HCO 3 - Acetoacetyl-CoA!-ketothiolase (Nmar_0841 or Nmar_1631) Acetoacetyl-CoA 3-Hydroxybutyryl-CoA dehydrogenase (Nmar_1028) Propionyl-CoA carboxylase (Nmar_0272, 0273, 0274) Methylmalonyl-CoA epimerase Methylmalonyl-CoA mutase (Nmar_0953, 0954, 0958) The 3OH-propionate/ 4 OH-butyrate cycle in Crenarchaeota 4-Hydroxybutyryl-CoA dehydratase (Nmar_0207) 4-Hydroxybutyryl-CoA 4-Hydroxybutyryl-CoA synthetase (Nmar_0206) Succinate semialdehyde reductase (Nmar_1110 or Nmar_0161) Succinate-semialdehyde Succinyl-CoA reductase (Nmar_1608)

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14 Photosynthetic organisms Distinction between light and dark reaction Light reaction conserve energy of light into chemical energy (ATP) Dark reaction involves the consumption of ATP for fixation of CO 2 Depending on electron donor: Oxygen-producing: oxygenic Non oxygen-producing: anoxigenic Oxygenic photosynthesis 2e - 2H + Water serves as electron donor

15 Anoxygenic photosynthesis Hydrogen sulfide (or sulfur) serves as electron donor

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17 Structure of a chloroplast Arrangement of light-harvesting Chlorophylls versus reaction center

18 Phylogenetic affiliation of phototrophic bacteria Oxigenic photosynthetic bacteria Cyanobacteria - Only bacteria which gain energy by oxigenic photosynthesis (formation of O 2 ) - Large and heterogeneous group of bacteria - Major primary producer in many habitats (aquatic and terrestrial habitats, symbiotic with Eukaryotes) - Ancestor of chloroplasts (Endosymbiosis theory) - Many can fix N 2 (Heterocyst or temporal seperation) - Occur as unicellular and filamentous forms

19 Absorption spectrum of cyanobacterium

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22 Phototrophic purple bacteria - gain energy by anoxic photosynthesis (no formation of O 2 ) - contain bacteriochlorophyll and a variety of carotonoids - electron carriers are arranged in specific intracytoplasmatic photosynthetic membranes (increase of pigment density) - electron carriers are in the order of more electronegative to higher electropositive reduction potential Intracytoplasmic membranes in anoxygenic phototrophs

23 Phototrophic purple sulfur bacteria Purple sulfur bacteria e.g. Chromatium okenii Gamma proteobacteria Habitat: stratified lakes Electron donor: reduced sulfur compounds H 2 S, S 0, S 2 O 3 2- Sulfur can be stores in globules inside the cell Mixotrophic: CO 2 fixation (Calvin Cyclus), organic acids Phototrophic purple sulfur bacteria Purple sulfur bacteria Ectothiorhodospira sp., Halorhodospira sp. Gamma proteobacteria Habitats: sola lakes, marine environments halophilic = salt-loving Produce sulfur outside the cell Electron donor: reduced sulfur compounds H 2 S, S 0, S 2 O 3 2- Mixotrophic: CO 2 fixation (Calvin Cyclus), organic compounds

24 Phototrophic purple nonsulfur bacteria e.g. Rhodospirillum rubrum Alpha or beta proteobacteria Electron donor: hydrogen, sulfur, organic substrates (no storage of sulfur) Some can grow in the dark by fermentation, anaerobic respiration, or aerobic respiration Can also fix N 2 Mixotrophic: CO 2 fixation (Calvin Cyclus), organic compounds

25 Vesicular photosymthetic membranes Rhodobacter capsulatus Anoxygenic photosynthesis in purple bacteria Only 1 light reaction!

26 Arrangement of protein complex in phototrophic purple bacteria Green sulfur bacteria z.b. Chlorobium limicola Phylum green sulfur bacteria All isolates are obligate anaerobic and phototrophic contain chlorosoms (location of photosynthesis) electron donors: reduced sulfur compounds H 2 S, S 0, S 2 O 3 2- produced sulfur resides outside the cells Mixotrophic: CO 2 fixation (reverse citric acid cycle) organic compounds (photoheterotrophy)

27 Chlorosomes in green sulfur bacteria Chlorophyl-rich bodies, connected to cytoplasma membrane Model of chlorosome structure (green sulfur and green nonsulfur bacteria)

28 Green Sulfur bacteria Consortia "Chlorochromatium aggregatum" Symbiosis between Phototrophic green sulfur bacterium (epibiont) and a chemotrophic beta proteobacterium (by J. Overmann, mikrobiologischer-garten.de) Green nonsulfur bacteria ( Chloroflexi ) e.g. Chloroflexus aurantiacus All isolated members are thermophilic Formation of thick microbial mats in hot habitats. Electron donor: H 2 and organic compounds CO 2 fixation via 3-hydroxypropionate cycle Heterotrophic with organic acids In the dark, chemoorganotrophic by aerobic respiration

29 Green nonsulfur bacteria ( Chloroflexi ) Chloroflexus aurantiacus Heliobacteria z.b. Heliobacillus chlorum contain bacteriochlorophyl g! Strict anaerobic, N 2 -fixation! Anoxygenic phototrophic Gram-positive bacteria! Spore-forming Electron donor: H 2 and pyruvate (fermentation) Mixotrophic: CO 2 fixation (reverse citric acid cycle) organic compounds

30 Bundles of cells of Heliophilum fasciatum Spore formation Heliobacterium gestii

31 Physiological properties of phototrophic Bacteria Cyanobacteria Purplebacteria Green Sulfur bacteria Green non- Sulfur bacteria Heliobacter PS-type PS I and II PS II PS I PS II PS I Pigments Chl a (b) BChl a, b BChl a, c, (d, e) BChl a, c BCHl g Autotrophy + (+) + +/- -(?) Physiology Photoauto- Lithoauto- Photoauto- Lithoauto- Organohetero- Photoauto- Lithoauto- Photoauto- Lithoauto- Organohetero- Photoauto- Organohetero- CO 2 fixation Calvin-cycle Calvin-cycle Reductive TCA 3OH-Propionate None? Electron donor H 2 O H 2 S/ organic H 2 S H 2 / organic Organic Adapted from Fuchs and Schlegel Allgemeine Mikrobiologie Comparison of electron flow