Amyris Raises Additional US$ 70 Million for Micriobial Biofuels Production

Amyris Biotechnologies today announced the first portion of their B round financing for US$ 70 million.  This brings the total company financing for microbial production of biofuels to just under US$ 100 million in the last year.  The press release also notes Amyris already has bugs in the lab producing "bio-jet", "bio-diesel", and "bio-gasoline".  The latter is interesting because previous announcements had suggested butanol as a target product rather than a hydrocarbon.  Immiscible hydrocarbons will be much easier (read "less expensive") to separate from the fermentation broth than water soluble alcohols.

In any event, the company is clearly moving faster than even my earlier optimistic estimates (see "The Need for Fuels Produced Using Synthetic Biology").  While the speed of engineering efforts is still an issue (see "The Intersection of Biofuels and Synthetic Biology"), and will be for some time to come, I have been spending more time lately trying to understand the issue of scale.  The petroleum industry is absolutely enormous, and replacing any significant amount of petro-fuels with bio-fuels will require feedstocks in abundance.  It is by no means clear that the U.S. can meet the demand with domestic biomass production.  More on this as the topic develops.

Sony's Enzyme-Powered, Sugar Fueled Power Supply

Sony has apparently demonstrated a power supply for consumer electronics that uses enzymes to covert sugar to useful electrons (via Gizmodo).  Not many details are available (to non-Japanese speakers, anyway), but it looks like each "module" generates ~50 mW from an unspecified amount of sugar.  It is evidently just an engineering demonstration, but it's pretty cool nonetheless.  No word on how the digested sugar is converted to electrical power.

The Intersection of Biofuels and Synthetic Biology

New players are appearing every day in the rush to production biofuels using synthetic biology.  I just noticed an announcement that Codon Devices has signed an agreement with Agrivida for:

The discovery, development, and commercialization of engineeredproteins for use in so-called 'third generation' biofuel applications. Under the terms of this agreement, Codon Devices will deliver to Agrivida optimized enzymes to be embedded in crops for biofuels production.

...Agrivida, an agricultural biotechnology company, is developing such third generation biofuels by creating corn varieties optimized for producing ethanol. First generation methods for manufacturing ethanol make use of the corn grain only, leaving the remaining plant material, such as the corn leaves, stalks, and husks in the field. Central to Agrivida’s ethanol-optimized corn technology are engineered cellulase enzymes that are incorporated into the corn plants themselves. These enzymes will efficiently degrage the entire mass of plant material into small sugars that can then be readily converted to ethanol.

The step of putting some of the biofuel processing into crops was inevitable, but I can't say I am particularly thrilled about it.  I am not opposed to the principle of open planting of GM crops, but, because many GM plants do not behave as predicted once placed in a complex ecosystem (i.e., nature), I wonder if  we shouldn't be more circumspect about this particular engineering advance.

In other interesting developments at Codon, they also recently announced a deal with Open Biosystems wherein the latter will:

Sell and distribute Codon Devices’ gene synthesis offering to researchers with needs that fall below Codon’s minimum order threshold.  The partnership will enable a wide range of new customers to utilize high-quality, low-cost gene synthesis in their research, and will greatly strengthen Codon Devices’ presence within academic, government and other non-profit institutions.

I also notice Codon is now advertising gene synthesis for $.69 per base for constructs between 50 and 2000 bases in length, with "typical delivery" in 10-15 days.  2001-5000 bases will cost you $.84 per base and 15-20 days.  Last year at SB 2.0, Brian Baynes suggested they would be at about $.50 per base within a year, so costs continue to fall pretty much apace.  But delivery times are staying above two weeks, and this is now becoming a problem for some of Codon's customers.  I am not at liberty to divulge names, but some synthetic biology companies that rely on outside gene synthesis are starting to chafe at having to wait two weeks before trying out new designs.  This is something we predicted would happen in the "Genome Synthesis and Design Futures" report from Bio-era, though I am a bit surprised it is happening so soon.  This may be another indication of how quickly SB is becoming an important technology in the economy.  Engineers trying to turn around products aren't satisfied with the NIH/academic model of trading off time for money -- the market, to first order, only cares about products that are actually for sale, which means those that make it through R&D quickly and generate revenues in what will become an increasingly crowded field.

Concerns about delays in the R&D cycle due to outsourced gene delivery are also becoming confounded by IP issues.  Personally, I am certainly not thrilled about sending my protein designs around via email, and I know of another SB company (which again I am not at liberty to name) that is becoming less and less comfortable with sending sequences for new genetic circuits out the door in electronic form.  This can only be exacerbated by the deal Codon Devices has just signed with Agrivida, an explicit competitor to anybody trying to produce anything in hacked/engineered organisms.  A couple of months ago, I had a conversation with Brian Baynes (which I will post here sometime soon) in which he outlined Codon's plans for participating in markets beyond gene synthesis.  I suspect Codon Devices will have to start paying more and more attention to conflict of interest issues generated by its simultaneous role as a fabrication house and provider of design services.

I'll argue again that the two trends of IP concerns and R&D time scales will drive the emergence of a market in desktop gene synthesis machines, whether you call them "desktop gene printers" or  something else.  This weekend at SciFoo, Drew Endy suggested such instruments are a long ways off.  Drew has been paying more attention to the specific engineering details of this than I have, if for no other reason that his involvement in Codon, but, in addition to my own work, I think that there are enough technological bits and pieces already demonstrated in the literature that we could see a desktop instrument sooner rather than later; that is, if a market truly exists. 

Environmental Effects of Growing Energy Crops

News this week that the dead zone in the Gulf of Mexico, caused by agricultural run-off from the mid-west, is again this year going to be quite large.

There is some disagreement about exactly how large.  A article from Minnesota Public Radio leads off with: "A scientist with the National Oceanic and Atmospheric Administration, NOAA, says this summer's dead zone could be as large as 8500 square miles. That's 77 percent larger than the average size of the dead zone over the last two decades."

The article continues:

The issue of nitrogen is especially important this year because it's the main fertilizer used on the nation's corn crop.

U.S. farmers this spring planted one of their largest corn crops ever, up almost 20 percent from a year ago. Much of the increase will go to meet the demands of the ethanol industry.

Runoff from farm fields carries nitrogen into streams and rivers and eventually the Gulf of Mexico. NOAA's David Whitall says the corn-biofuels-dead zone link is one area researchers will examine as they search for answers.

...One federal study says if ethanol production continues to expand, nitrogen loads to the Gulf could increase another 30 percent.

At CNN, on the other hand, the size of the dead zone is portrayed somewhat differently:

The oxygen-poor "dead zone" off the Louisiana and Texas coasts isn't quite as big as predicted this year, but it is still the third-largest ever mapped, a scientist said Saturday.

...The 7,900-square-mile area with almost no oxygen, a condition called hypoxia, is about the size of Connecticut and Delaware together. The Louisiana-Texas dead zone is the world's second-largest hypoxic area, she said.

This year's is about 7.5 percent smaller than [had been] predicted, judging by nitrogen content in the Mississippi River watershed.

[Previous predictions were] about 8,540 square miles, which would have made it the largest measured in at least 22 years. More storms than normal may have reduced hypoxia by keeping the waters roiled.

No mention at CNN of any role any role in the dead zone of biofuels.  The difference between the numbers cited by the two sources is less than 5%, which probably isn't a big deal, especially give then fact that neither article cites error bars.  But there is a difference in focus.  On the one hand, the dead zone is bigger than ever, on the other, not so bad.  Corn acres are certainly up in the U.S., and the effects of the consequent increase in irrigation and fertilizer use is something to keep an eye on.

LS9 - "The Renewable Petroleum Company" - in the News

LS9, "The Renewable Petroleum" Company, has just hired a former oil executive as its new CEO.  The promise of direct microbial fuel production is so great that this news even made The Huffington Post.  Why all the sudden buzz?  The answer is that this technology is really quite new, but is making great strides.  Moreover, as I wrote about a couple of weeks ago (see "The Need for Fuels Produced Using Synthetic Biology"), the economics of producing fuels from microbes is so radically different from what we are used to that it will upend our notions industrial infrastructure.  That said, it will still take some time before all the impacts are fully appreciated.

Last Thursday, I did a short interview for the series Questions for the Future, produced by CNBC Europe/Asia in association with Shell, during which the host was somewhat perplexed about why there was not yet more widespread discussion of this technology.  At The Huffington Post, David Roberts starts his post with a note of skepticism:

Picture a liquid fuel that is derived from the same feedstocks as cellulosic ethanol (switchgrass, sugar cane, corn stover) but contains 50% more energetic content and is made via a process that uses 65% less energy.

Unlike cellulosic ethanol, this fuel can be distributed via existing oil pipelines rather than gas-hogging trucks and trains, dispensed through existing gas stations rather than specialized pumps, and used in existing engines rather than modified "flex-fuel" engines.

In short, it is a biofuel that can be substituted directly and immediately for gas or diesel, on a gallon-for-gallon basis.

Sounds pretty good, eh? Too good to be true?

Which illustrates one reason why this topic isn't so much in the news.  It does sound too good to be true.  But it is quite real, with Amyris Biotechnologies on track to produce jet fuel from microbes by 2011 at an equivalent cost of US$ 40 per barrel.

Another interesting thread to this discussion is the potential internal conflict generated in "Greens" by the notion of reducing carbon emissions ("Good!") using genetically modified organisms ("Bad!").  I've been working this idea into an essay about laying the foundations for a bio-economy, but Roberts makes it explicit in his post; "I know there are greens who feel creepy about genetic engineering, and they probably won't like the fact that LS9 is trying to patent a life form. But I don't really share those concerns, so I'll just skip them."  No worries.  Just like that.  I am not so certain Greenpeace et al. will follow along so quietly.

In the press release from LS9, new CEO Robert Walsh says:

After years of leadership roles in the traditional petroleum industry and responsibility over all aspects of the hydrocarbon supply chain, I can see clearly how LS9's products will fit into existing infrastructure and deliver significant value to partners and consumers compared with other biofuel alternatives.  LS9 has the opportunity to fundamentally change the transportation fuel equation, which makes me incredibly excited to join this talented team.

While it's true that these engineered synthetic fuels will likely find first use within existing distribution channels, it is the potential for distributed manufacturing that truly changes the game.  It will be interesting to see how long it takes for this part of the story to work its way into the broader conversation.

Finally, here is additional coverage of the LS9 announcement at GreenCarCongress.

European Biofuels Travelblog

Instead of "Cellular Lipo-Sculpting Eye Gel", this time the transcontinental party favor was "Relaxing Yuan Zhi Pulse Point Balm".  Thank you British Airways.  The chocolate mousse was mighty tasty, though.  Almost as good as on Air France.  Almost.

Thankfully, the continental leg of the trip was canceled.  Due to security at Heathrow and intense weather across the U.K., I can't say I was disappointed.  Audiences in London and Edinburgh were at least as attentive as those we presented to in Asia, and I suspect this due in part to real concerns about carbon emissions throughout Europe.  In Asia, biofuels seemed to be thought of as more a business opportunity, with carbon emissions as a complication.  In the U.K., in addition to the basic economic concerns, the questions about carbon were more along the lines of trying to understand which fuels and which technologies actually reduce emissions.

Crazy moment of the trip: who should we bump into in the lobby of our hotel in London but John Melo, CEO of Amyris Biotechnologies.  Small world.

More Amyris and Biofuels News

Amyris Biotechnologies today announced new additions to their Board of Directors, all of whom are formerly associated with BP.  As I have suggested before, Amyris is presently the company to watch in the race for direct microbial production of biofuels (see "The Need for Biofuels Produced Using Synthetic Biology").

Note that, in the "About Amyris Biotechnologies" blurb at the end of the release, biofuels now dominate the stated goals of the company.  I wonder if this might eventually squeeze out the original focus on inexpensive malaria drugs:

   Amyris Biotechnologies (www.amyrisbiotech.com) combines break-through technology and unique insights in the transportation fuels sector to bring environmentally friendly fuels to market. Amyris believes its microbial technology will allow it to reduce the production cost of artemisinin-based anti-malarial treatment to a fraction of its current cost. Amyris is leveraging its technology platform to provide a cost-competitive bio-gasoline, a bio-diesel, and a bio-jet solution that works in current engines and distribution infrastructure without compromising fuel performance. All Amyris biofuels are designed to provide consumers and end users uncompromising alternatives to petroleum-based fuels. Amyris is a privately-held venture-backed company whose investors include Khosla Ventures, Kleiner Perkins Caufield & Byers (KPCB), and TPG Ventures.

The Need for Fuels Produced Using Synthetic Biology

Among the most promising short term applications of Synthetic Biology is biological production of liquid fuels.  But beyond the technical and economic attraction of the project, the reasons we require progress in this area are manifest; diversification of fuel sources thereby reducing dependency on imports, improving air quality, reducing greenhouse gas and particulate emissions that contribute to climate change, eliminating the present coupling between biofuels and food crops, and carbon sequestration.

Bio-era is in the middle of scheduled briefings in Asia, the U.S., and Europe describing the present state of biofuels markets and associated technologies, and these trips, along with recent headlines concerning commodities prices and future fuel demands, have helped clarify the story in my mind.  Below I outline some of the factors in play:

Carbon and other Greenhouse Gas Emissions: The amount of water coming off Antarctica and Greenland scares the crap out of me.  It's true that this isn't my professional specialty, but I have been following the literature on polar ice mass and movement for a decade.  The news is just getting worse.

The present coupling between biofuels and food crops creates upward pressure on food price inflation and reduces (or eliminates) the economic incentive to produce biofuels: Ethanol demand has pushed up the price of corn, and in the U.S. politically motivated trade barriers to Brazilian ethanol derived from sugar cane threaten to keep corn prices high.  Palm oil is presently trading at historic highs, and at a ~30-40% premium to finished diesel, but this is actually driven by food demand, primarily from India and China.  I am a simple physicist by training, rather than a sophisticated economist, but given the increase in food demand I don't see the price coming down even with increased supply.  This puts anybody planning to refine palm oil into biodiesel completely underwater for the foreseeable future. 

China (and India) will require increasing resources over the coming decades: More on this in posts to come.  The numbers are mind boggling.

Ethanol is by no means an advanced biofuel; from both a technical and an economic perspective ethanol is a backwards biofuel: The future is all about producing biofuels that are high energy content (not ethanol), are not water soluble (not ethanol), can be easily integrated into the existing gasoline and diesel distribution infrastructure (not ethanol), and require minimal, if any, initial changes in engine technology (not ethanol).  The average age of an automobile in the U.S. is now at least 10 years (depending on who is counting, and how), which means engine technology turns over very slowly here.  It is faster in other countries (2-3 years in Japan, if memory serves), but this dramatically influences the speed with which new fuels can enter the market.

You don't want to be long on petroleum in ten years:

First, despite a greater than 10% annual growth in auto sales in China, petroleum demand has evidently plateaued due to increased biofuel blending.  I'm not sure I completely believe this yet, but it is an interesting assertion.

Second, three companies are already out in front with funding to use both traditional metabolic engineering and synthetic biology to produce microbes that churn out biofuels:

LS9 is "Developing Renewable Petroleum biofuels: new, clean, and sustainable fuels that fulfill our long and short term energy needs. Derived from diverse agricultural feedstocks, these high energy liquid fuels are renewable and compatible with current distribution and consumer infrastructure."

Synthetic Genomics, Craig Venter's shop, just announced a partnership with BP aimed at using organisms and genes found in subsurface hydrocarbon deposits to develop "cleaner energy production and improved recovery rates".

Amyris Biotechnologies recently received $20 million to develop direct microbial production of liquid biofuels.  Amyris, in particular, is well positioned to make some serious headway.  The company website suggests they are well on their way to making both butanol and biodiesel  (or more likely a precursor to diesel?) in microbes.  In an article in Technology Review, the new CEO, John Melo, says the company has already developed a metabolic pathway to produce a fuel equivalent to Jet-A.  This is particularly interesting given the recent announcement by the U.S. Air Force that it will replace at least 50% of its petroleum use with synthetic fuels by 2010.  In an article by Don Phillips, The New York Times is reporting that, "The United States Air Force has decided to push development of a new type of fuel to power its bombers and fighters, mixing conventional jet fuel with fuels from nonpetroleum sources that could eventually limit military dependence on imported oil."  At the moment, the immediate plan appears to utilize a synthetic fuel produced using natural gas, but anybody who can crack the aviation biofuel nut has immediate access to a 3.2 billion gallon per year market in the Air Force alone.

So how long is this all going to take?  Amyris CEO Melo mentions they hope, "To make a Jet-A equivalent with better properties on energy and freezing point with a $40 barrel cost equivalent by 2010 or 2011".  That's faster than I was expecting, but I find the time scale highly credible.  Below is a figure with data drawn from Jay Keasling's recent presentation to the UC Berkeley faculty senate on BP's investment in the Energy  Biosciences Institute.

Isoprenoid_yieldThe data represents a roughly billion-fold improvement in yield over 6 years.  (I've called this "pre-synthetic biology improvements" because the data is the result of applying fairly traditional metabolic engineering techniques, rather than the combination of Biobricks.  This is by no means a critique of Jay Keasling or his teams at UC Berkeley or Amyris, but rather a simple contrast of methodology.)

You would be hard pressed to find examples of that magnitude of improvement in any human industrial process over any 6 year period, but that is exactly what is possible when you turn to biology.  Moreover, the complexity of the isoprenoid pathway is probably about the same as you would expect for producing biobutanol or a Jet-A equivalent.  This is why John Melo is bullish about making progress on biofuels.  Given that Amyris is evidently already on the path towards butanol, diesel, and aviation fuel, five years is by no means an overly optimistic estimate of reaching commercial viability.  Note that this doesn't mean Amyris takes over the liquid fuels market overnight.  It can take decades for new technologies to make progress against existing infrastructure and investment.

But assuming Amyris, or any other company, is successful in these projects, it is worth considering first the resulting impact on the liquid fuels market, then more generally the effects on structure of the economy as a whole.

The economic considerations of scaling up direct microbial producing of biofuels are fundamentally and radically different than those of traditional petroleum production and refining.  The costs associated with finding a new oil field and bringing it into full production are considerable, but are also so variable, depending on location, quality, and local government stability, that they are a poor metric.  But a very clean measure of increasing gasoline and diesel supplies is the fractional cost of adding refining capacity, presently somewhere between US$ 1 and 10 billion dollars for a new petro-cracking plant, plus the five or so years it takes for construction and tuning the facility for maximum throughput.

In contrast, the incremental cost of doubling direct microbial production of a biofuel is more akin to setting up a brewery, or at worst case a pharmaceutical grade cell culture facility, which puts the cost between about US$ 10,000 and 100,000,000.  Pinning down the cost of a biofuel production facility is presently an exercise in educated speculation, but it is more likely to be on the low end of the scale suggested above, particularly for a fuel like butanol, which, unlike, ethanol, is not soluble in water and therefore does not require distillation; it can simply be pumped or skimmed off the top of the tank in a continuous process.  Beer brewing presently occurs at scales from garage operations bottling a few liters at a time to commercial operations running fermenters processing thousands to many millions of liters per year.  Thus, once in possession of the relevant strain of microbe, increasing production of a biofuel may well be feasible at the local level, thereby matched to fluctuations in demand.  Microbial biofuels could therefore be an excellent initial demonstration of distributed biological production (PDF warning).

In the end, the scalability of microbial production of biofuels depends in part on what materials are used as feedstocks, where those feedstocks come from, and how they are delivered to the site of production.  Whereas petroleum products are a primary feedstock of today’s economy, both as a raw material for fabrication and for the energy they deliver, it may eventually be possible to treat biomass or waste material as feedstocks for microbes producing more than just fuels.  But as I observed above, any biological production process  for biofuels that relies on a sugar or starch crop also used in food production will be subject to the same skewed market dynamics now playing out between food and conventional biofuels.

There are clear challenges to overcome in the years ahead, but given the progress already demonstrated I am comfortable we will find solutions with continued effort.

A Synthetic Enzymatic Pathway for Hydrogen Production from Starch

Zhang, et al., demonstrate a synthetic pathway consisting of 13 enzymes that turns starch into hydrogen.  The paper, at PLoSone, makes clear how important it is to ensure that the definition of Synthetic Biology, if there is a definition yet, includes not just new circuits in cells and new organisms but also cell free systems.  The article (High-Yield Hydrogen Production from Starch and Water by a Synthetic Enzymatic Pathway) is open access, so I won't bother to quote from it extensively.

But the details are pretty cool.  The authors used 11 off-the-shelf enzymes (from spinach, rabbit, E. coli, and yeast) ordered from Sigma, and two they purified themselves (from coli, and P. furiosus).  I imagine it won't be too long before those last two are also available commercially. Here is a paragraph that sums up the context of what the authors accomplished and where they will look for performance improvements:

This robust synthetic enzymatic pathway that does not function in nature was assembled by 12 mesophilic enzymes from animal, plant, bacterial, and yeast sources, plus an archaeal hyperthermophilic hydrogenase. The performance (e.g., reaction rate and enzyme stability) is anticipated to be improved by several orders of magnitude by using the combination of (a) enzyme component optimization via metabolic engineering modeling, (b) interchangeable substitution of mesophilic enzymes by recombinant thermophilic or even hyperthermophilic enzymes, (c) protein engineering technologies, and (d) higher concentrations of enzymes and substrates. ... This research approach will naturally benefit from on-going improvements by others in synthetic biology systems that are addressing cofactor stability, enzyme stability by additives, and co-immobilization, and development of minimal microorganisms that can be built upon to create an in vivo enzyme system that produces H2 in high yields.

What I think is most interesting about all this is that Zhang and colleagues have effectively just put a whole bunch of new Biobricks on the table.  Moreover, those new parts for producing biofuels are reasonably well characterized, at least in vitro.  A cursory search of the Registry of Standard Biological Parts doesn't turn up any of these enzymes, but since the gene sequences are either already in Genbank or are relatively easy to generate, this gap points to an area of expansion for Biobricks in general and the International Genetically Engineered Machines Competition (iGEM) in particular.

I suggest it is time to expand the horizons of iGEM beyond demonstration projects to start tackling real world problems.  The student teams have made fantastic progress over the past few years, with some projects winding up on the cover of high-profile journals.  I would like to see this year's iGEM participants take on biological production of fuels and materials, bioremediation, and biological carbon sequestration.  Even if popular attention has yet to come round, the problems we presently face are enormous (see my post "It's Time to Invest in Water Wings"), and through the combination of enthusiasm and creativity iGEM participants could start developing solutions today.