ARPA-E Adams-Kelly 10-21-09...Chalcopyrite, t = 0 days with M. sedula Chalcopyrite, t = 21 days with...
Transcript of ARPA-E Adams-Kelly 10-21-09...Chalcopyrite, t = 0 days with M. sedula Chalcopyrite, t = 21 days with...
H2-driven CO2 Fixation by Extremely Thermoacidophilic Archaea
Michael W.W. Adams, University of Georgia
Robert M. Kelly, North Carolina State University
ARPA-E Workshop on Direct Solar FuelsOctober 21, 2009
LIGHT DARK
H2O H2
ElectronSource
PrimaryReducedProduct
Biofuel
CH/O
CO2
ElectronSource
PrimaryReducedProduct
Biofuel
SOLAR BIOFUEL PRODUCTION
OxidizedElectronSourceO2
SecondaryReducedProductSUGARS
OxidizedSubstrate
CO2
ElectronSourceH2O
PrimaryReducedProductNADPH
SUGARS
Green Plants
CO2
VISIBLE LIGHT
Biofuel(of choice*)
BiomassConversion
MicrobialConversion
PS CC
PS = photosynthesis (I+II)CF = CO2 fixation (Calvin cycle)
ElectronSource
PrimaryReducedProduct
Biofuel
SOLAR BIOFUEL PRODUCTION
*CxHy / CaHbOc
OxidizedElectronSourceO2
SecondaryReducedProductSUGARS
OxidizedSubstrate
CO2
ElectronSourceH2O
PrimaryReducedProductNADPH
Algae
CO2
VISIBLE LIGHT
Biofuel
H2
PS CC
PS = photosynthesis (I+II)CF = CO2 fixation (Calvin cycle)
ElectronSource
PrimaryReducedProduct
Biofuel
SOLAR BIOFUEL PRODUCTION
OxidizedElectronSourceO2
SecondaryReducedProductSUGARS
OxidizedSubstrate
CO2
ElectronSourceH2O
PrimaryReducedProductNADPH
CO2
VISIBLE LIGHT
MetabolicEngineering Biofuel
(of choice)
Algae
Simplified (Biomimetic)Scheme?
PS CC
PS = photosynthesis (I+II)CF = CO2 fixation (Calvin cycle)
ElectronSource
PrimaryReducedProduct
Biofuel
SOLAR BIOFUEL PRODUCTION
OxidizedElectronSourceO2
OxidizedSubstrate
CO2
ElectronSourceH2O
PrimaryReducedProduct
H2
CO2
VISIBLE LIGHT
Biofuel(of choice)
PS = photosynthesis (I+II)CF = CO2 fixation (Calvin cycle)
CFModule
PSModule
ElectronSource
PrimaryReducedProduct
Biofuel
SOLAR BIOFUEL PRODUCTION
OxidizedElectronSourceO2
OxidizedSubstrate
CO2
ElectronSourceH2O
PrimaryReducedProduct
H2
CO2
VISIBLE LIGHT
Biofuel(of choice)
PS = photosynthesis (I+II)CF = CO2 fixation (Calvin cycle)
O2
CO2 + H2O
Fuel use withoxidized source(O2) regenerates
substrates (CO2, H2O)
IDEAL SOLARENERGY CONVERSION
SYSTEM
ElectronSource
PrimaryReducedProduct
Biofuel
SOLAR BIOFUEL PRODUCTION
CFModule
PSModule
OxidizedElectronSourceO2
OxidizedSubstrate
CO2
ElectronSourceH2O
PrimaryReducedProduct
H2
CO2
VISIBLE LIGHT
Biofuel(of choice)
PS = photosynthesis (I+II)CF = CO2 fixation (Calvin cycle)
O2
CO2 + H2O
Fuel use withoxidized source(O2) regenerates
substrates (CO2, H2O)
IDEAL SOLARENERGY CONVERSION
SYSTEM
LIGHT DARK
H2O H2
ElectronSource
PrimaryReducedProduct
Biofuel
SOLAR BIOFUEL PRODUCTION
CH/O
CFModule
PSModule
CO2
PSM CFM
OxidizedElectronSourceO2
OxidizedSubstrate
CO2
ElectronSourceH2O
PrimaryReducedProduct
H2
CO2
VISIBLE LIGHT
Biofuel(of choice)
PS = photosynthesis (I+II)CF = CO2 fixation
Key Research
Areas
PS
LIGHT DARK
H2O H2
ElectronSource
PrimaryReducedProduct
Biofuel
SOLAR BIOFUEL PRODUCTION
CH/OPSM CFM
CFModule
PSModule
CO2
CO2 CHO/CH
2H+ + 2e- H2
H2
H2
OxidizedElectronSourceO2
OxidizedSubstrate
CO2
ElectronSourceH2O
PrimaryReducedProduct
H2
CO2
VISIBLE LIGHT
Biofuel(of choice)
PS = photosynthesis (I+II)CF = CO2 fixation
Key Research
Areas
PS
LIGHT DARK
H2O H2
ElectronSource
PrimaryReducedProduct
Biofuel
SOLAR BIOFUEL PRODUCTION
CH/OPSM CFM
CFModule
PSModule
CO2
CO2 CHO/CH
2H+ + 2e- H2
H2
Key Biological ReactionsPSM:1. Water photolysis (visible light):
H2O O2 + 2H+ + 2e-
2. Hydrogen production2H+ + 2e- H2
CFM:1. Hydrogen activation
H2 2H+ + 2e-
2. CO2 to biofuel conversion (reduced carbon/CH/CHO) CO2 + 2H+ + 2e- CxHy / CaHbOc
Much higher energy yield using H2 as reductant for CO2 fixation to biofuel compared to sugar to biofuel
LIGHT DARK
H2O H2
ElectronSource
PrimaryReducedProduct
Biofuel
SOLAR BIOFUEL PRODUCTION
CH/OPSM CFM
CO2
Structure/Function Studies of NiFe-hydrogenaseStructure/Function Studies of NiFe-hydrogenase
- Hydrogenase is not a ‘commodity enzyme’
- Very complex maturation process in vivo to assemble complex, O2-sensitive, NiFe-catalytic site (CO, CN ligands, etc.)
- No genetically-tractable organism that produces hydrogenase available for detailed structure/function studies
- Recent breakthrough – production of recombinant form of the NADPH-dependent Pyrococcus hydrogenase in E. coli
Fundamental structure/function studies to design:
- ‘Minimal’ hydrogenases(NiFe-site plus peptides)
- H2 production and H2 activation catalysts
- Decreased O2-sensitivity
- Electron carrier specificity
- Pathways of electron transfer
Hydrogen Production from Polyglucose(13 enzymes, Pi, NADP – catalytic)
(C6H10O5)n + 7 H2O 12 H2 + 6 CO2
PPP(Pentose
PhosphatePathway)
#13: NADPH-dependentH2-evolving
Pyrococcus Hydrogenase
Zhang et al. PLoS One 5, e456, 2008
Calvin Cycle
Reverse Tricarboxylic Acid Cycle Reductive Acetyl-CoA Pathway
Adapted from http://lecturer.ukdw.ac.id/dhira/Metabolism/CarbonAssim.html
plants, cyanobacteria, purple and green bacteria
Chlorobium limicola
Desulfobacter
hydrogenophilus
Hydrogenobacter thermophilus
Acetobacterium woodii
Defulfobacterium autotrophicum
Microbial CO2Fixation Pathways
Dicarboxylate/4-Hydroxybutyrate Cycle 3-Hydroxypropionate/4-Hydroxybutyrate Cycle
Ramos-Vera, W. H. et al. (2009) Berg, I. A. et al. (2007)
3-Hydroxypropionate Cycle
Chloroflexus aurantiacusAlber, B. et al. (2006)
Ignicoccus hospitalis Metallosphaera sedula
Microbial CO2Fixation Pathways
(Thermophiles)
The Extremely Thermoacidophilic Archaeon Metallosphaera sedula
The Extremely Thermoacidophilic Archaeon Metallosphaera sedula
M. sedula genes closest sequence matches
S. solfataricus
S. tokodaii
S. acidocaldarius
Other sulfolobus species(islandicus, metallicus, shibatae)Acidianus species
Ferroplasma species
Thermoplasma species
Pyrobaculum species
Other / No top hit
SsoSac
Sto
other
Huber et al. 1989
Auernik, K. S., Y. Maezato, P. H. Blum, and R. M. Kelly. 2008. The genome sequence of the metal-mobilizing, extremely thermoacidophilic archaeon Metallosphaera sedula provides insights into bioleaching-associated metabolism. Appl. Environ. Microbiol. 74:682-92.
Genome: 2.2 Mb (46.2% G + C) 2304 ORFsGrowth: Mixotrophic aerobe; metal mobilizer
Topt = 73°C; pHopt = 2.0
Comparative genomics: Extreme thermoacidophilesComparative genomics: Extreme thermoacidophiles
Comparative genomics: Extreme thermoacidophilesComparative genomics: Extreme thermoacidophiles
Growing E. coliGrowing E. coli
Oxoid
Metal mobilization by M. sedulaChalcopyrite, t = 0 days with M. sedula Chalcopyrite, t = 21 days with M. sedula
CuFeS2 solids10g/LpH 2
Heavy metal tolerance
Iron oxidation
Sulfur oxidation
(Inorganic) Carbon fixation
Adhesion to solids
Cu2+
Auernik, KS, Y. Maezato, PH Blum, and RM Kelly. 2008. Appl. Environ. Microbiol. 74(3): 682-692Auernik, KS, CR Cooper, and RM Kelly. 2008. Curr. Opin. Biotechnol. 19(5):445-453
Auernik, KS and RM Kelly. 2008. Appl. Environ. Microbiol. 74(24): 7723-7732
Metal and RISC oxidation components on Fe2+, S°
Metal and RISC oxidation components on CuFeS2
Auernik, KS, and RM Kelly. Molecular hydrogen impacts chalcopyrite bioleaching efficiency for the extremely thermoacidophilic archaeon Metallosphaera sedula. (submitted)
Versatile growth physiology of M. sedula
Auernik, KS, and RM Kelly. Physiological versatility of the extremely thermoacidophilic archaeon Metallosphaera sedulasupported by heterotrophy, autotrophy and mixotrophy transcriptomes (submitted)
Autotrophy (12 hrs)
Heterotrophy (5 hrs)Mixotrophy (< 4hrs)
M. sedula Hydrogenases
Auernik, KS, and RM Kelly. Physiological versatility of the extremely thermoacidophilic archaeon Metallosphaera sedulasupported by heterotrophy, autotrophy and mixotrophy transcriptomes (submitted)
Auernik et al. (2008) Curr. Opin. Biotech. 19, 445-453
ATP
2 NADPH
NADPHATP
ATP
ATP
NAD
2 NADPH
M. sedula 3-HP/4-HB CO2 fixation pathway
Overall: 2CO2 + 4ATP + 5NADPH + CoA + NAD Acetyl CoA + 3ADP+AMP + 3Pi + PPi + 5NADP + NADH
Auernik, KS, CR Cooper, and RM Kelly. 2008. Curr. Opin. Biotechnol. 19(5):445-453 Berg, I et al. 2007. Science. 318:1782-1786
H1H2 A1A2 M1M2
M. sedula 3-HP/4-HB CO2 fixation pathway
4-hydroxybutyryl-CoA dehydratase
Auernik, KS, and RM Kelly. Physiological versatility of the extremely thermoacidophilic archaeon Metallosphaera sedulasupported by heterotrophy, autotrophy and mixotrophy transcriptomes (submitted)
M. sedula 3-HP/4-HB CO2 fixation pathway
3-HP/4-HB CO2 Fixation Pathway in Metallosphaera sedula
Fundamental Studies Needed:
1. Photosystems I and II
2. Hydrogenases
3. CO2 fixation pathways
Key issues:
1. Mechanisms of catalysis and design of biomimetic systems?
2. Electron transfer pathways between the three systems?
LIGHT DARK
H2O H2
ElectronSource
PrimaryReducedProduct
Biofuel
SOLAR BIOFUEL PRODUCTION
CH/OPSM CFM
CO2