Synthetic Biology in the Quest for Renewable Energy Jay Keasling Berkeley Center for Synthetic Biology University of California & Lawrence Berkeley National Laboratory Berkeley, CA 94720
Jan 16, 2016
Synthetic Biology in the Quest for Renewable Energy
Jay Keasling
Berkeley Center for Synthetic BiologyUniversity of California &
Lawrence Berkeley National LaboratoryBerkeley, CA 94720
The need for renewable energy
• US Energy demands to grow
• Reduction of US CO2 emissions
• Production of clean, cheap energy
1990: 12 TW 2050: 28 TW
Renewable
Biomass: a source for renewable energy
• About half of the carbonaceous compounds in terrestrial biomass are cellulose.
• The net primary production of biomass is estimated to be 60 Gt/yr of carbon in terrestrial and 53 Gt/yr in marine ecosystems.
• Almost all of the biomass produced is mineralized again by enzymes which are provided by microorganisms.
• Polysaccharide hydrolysis is one of the most important enzymatic processes on earth.
Lignocellulose
• Nearly universal component of biomass
• Consists of three types of polymers:– Cellulose– Hemicellulose– Lignin
• All three are degraded by bacteria and fungi Component Percent Dry Weight
Cellulose 40-60%
Hemicellulose 20-40%
Lignin 10-25%
Cellulose
• Cellulose is a chemically homogeneous linear polymer of up to 10,000 D-glucose molecules, which are connected by ß-1,4-bonds.
Taken from http://www.lsbu.ac.uk/water/hycel.html
3-D Cellulose Structure
Hemicellulose
• Hemicellulose is a polysaccharide composed of a variety of sugars including xylose, arabinose, mannose.
• Hemicellulose that is primarily xylose or arabinose are referred to as xyloglucans or arabinoglucans, respectively.
• Hemicellulose molecules are often branched.
• Hemicellulose molecules are very hydrophilic.
• They become highly hydrated and form gels.
Hemicellulose structure
Cellulose to ethanol
Taken from Demain et al. 2005. Microbiol. Mol. Biol. Rev. 69:124-154.
Cellulose
Hemicellulose
Cellobiose
XyloseXylobiose
C. thermocellum
C. thermosaccharolyticum
Ethanol
Lactate
Acetate
Cellulase
Hemicellulase
60ºC
Cellulosome structure
Cellulosome structure
• Stable & flexible • Many subunits• Organization promotes synergistic action • Non-catalytic, multipurpose subunit
which is the core of cellulosome structure
• Scaffoldin - 1,800 amino acids; single Cellulose Binding Domain; Cohesins; anchors cellulosome to cell surface
Cellulosome structure
• More active against crystalline than amorphous cellulose
• Form lengthened corridors between cell & substrate
• Cellulose degradation aided by noncellulosomal cellulases & cellulosomes released into environment
Problems
• Products other than ethanol or hydrogen are produced from cellulose.
• Clostridia are difficult to engineer.
• Cellulosome is extremely complex making its transplantation to another microbe a significant hurdle.
Goal
• Improve yield of energy-rich molecules from cellulose– Engineer the cellulosome into a
genetically tractable microorganism (e.g., Bacillus subtilis)
– Develop clostridium genetics to the point that extraneous metabolic reactions can be eliminated
Synthetic Biology
• De novo design of biological entities– Enzymes– Biomaterials– Metabolic pathways– Genetic control systems– Signal transduction pathways
• Need the ability to write a ‘blueprint’
Why do we need synthetic biology?
• Synthesis of drugs or other molecules not found in nature– Designer enzymes– Designer cells with designer enzymes or existing
enzymes
Why do we need synthetic biology?
• Energy production– Production of hydrogen or ethanol– Efficient conversion of waste into energy– Conversion of sunlight into hydrogen
Why now?
• Advances in computing power
• Genomic sequencing• Crystal structures of
proteins• High through-put
technologies• Biological databases• Diverse biological
sampling/collection
Why here?
• LBL has played a central role in the development of most of the technologies that will be essential for synthesizing new bacteria.
• Synthetic biology will leverage major LBL programs– Joint Genome Institute– Genomes-to-Life– Advanced Light Source– Molecular Foundry– NERSC
Building a Super H2 Producer
Complex Polysaccharides
H2
Specialty & CommodityChemicals Ethanol
Building a new chromosome based on genome sequences
Identification of minimal
gene set
Maximizing renewable
resource utilization
Specific aims
• Determine chromosomal design rules and construct the basic superstructure for an artificial chromosome for our host organism.
• Determine the minimal number of genes necessary for a viable, yet robust bacterium.
• Determine the components of the cellulose degrading machinery necessary for cellulose utilization.
Integration with LBNL Projects
• Joint Genome Institute – Cellulose degraders sequenced by JGI and
artificial chromosome sequencing.
• Genomes to Life– Transcript and protein profiling using GTL
facilities.
• Molecular Foundry– The cellulose degradation machinery as a
model molecular motor.
• Synthetic Biology– New initiative at LBNL and UCB.
Technical Challenges
• Engineering a completely new organism is a daunting task.
• The cellulose degrading machinery is an incredibly complicated molecular machine that will require significant characterization in its native host before it can be engineered into a new host.
Benefits to LBNL
• Establish a new initiative in synthetic biology.
• Establish a new program in hydrogen/ethanol production.
• Utilize large sequence database from JGI.