Scientists at the Energy Department’s National Renewable Energy Laboratory (NREL) have developed an enzyme that could change the economics of biofuel conversion by converting biomass to sugars up to 14 times faster and much cheaper than competing catalysts in today’s enzyme cocktails.

This enzyme is called CelA, a cellulase from the bacterium Caldicellulosiruptor bescii. C. bescii, producer of the remarkable CelA enzyme, that was first discovered in warm-water pools in the Valley of Geysers on Russia’s Kamchatka Peninsula in the 1990s. Russian scientists found the enzyme somewhat promising, but it wasn’t until the NREL researchers conducted a thorough analysis and added improvements that its remarkable potential was realized.

Being a from bacteria rather than a fungus enlarges the possibilities. As noted CelA can digest not one, but two of three major components in biomass: both cellulose and xylan leaving only the lignin behind. Each of the three polymers typically requires several types of enzymes to deconstruct them to soluble precursor molecules that can then be upgraded to ethanol, drop-in fuels, or chemicals.

Enzyme Activity Abating and Boring Cellulose Xylan.  Click image for more info.

Enzyme Activity Abating and Boring Cellulose Xylan. Click image for more info.

CelA works in two mechanical ways instead of one. It is an ablater, scraping the valuable material off the cell walls of the plants. And it is also a borer, digging deep into the wall to grab more of the digestible biomass. It is the only enzyme known to dig pits into biomass; others only ablate.

Of particular interest is it can operate at much higher temperatures than other enzymes. That’s important because high temperatures mean faster action. Also, because it can operate above the boiling point of alcohol, the alcohol is separated naturally, saving a costly distillation step in the conversion process – and the high temperatures kill many of the microorganisms that would otherwise interfere with the process.

Its looking like a major biofuel event.

As a comparison, the best commercially used enzyme converted sugars to a 30% extent over seven days. The CelA converted to double that extent. While it took the alternative enzyme seven days to achieve that conversion, CelA, with a small boost from an extra beta glucosidase, achieved the doubling in just about two days.

NREL Senior Scientist Roman Brunecky pointed out, “If you can achieve in one day what typically takes seven, you are saving the better part of a week of processing. And that can have a huge economic impact.”

CelA alone is four to five times faster at breaking down sugars than the enzymes in today’s cocktails. A more typical usage would be CelA combined with a beta glucosidase – the improvement that makes it 14 times faster.

If the enzyme continues to perform well in larger tests, it could help drive down the price of converting cellulose and, with it, the price of everything from jet fuel to ethanol, butanol, drop-in fuels, and numerous chemicals. NREL has filed for patent protection on the enzyme formulation and the improvements made to the unusual enzyme.

Conventional wisdom in the enzyme industry has it that the important discoveries have already been made: that another cellulase, Cel7A, was the key enzyme to be the backbone of all the commercial enzyme cocktails. NREL’s discovery not only reveals that there is an intriguing enzyme that can work several times faster, but it is also a reminder that nature still has secrets to uncover.

Using electron microscopy, the NREL researchers and their partners at the University of Georgia found that CelA not only ablates the cell wall of lignocellulosic biomass, but excavates cavities into the surface. CelA also worked faster on raw biomass than it did on biomass pre-treated with chemicals.

The researchers found that the size of the holes in the plant walls was about the same size as the enzymes themselves. The inability of the enzyme to digest a space on the surface that is any bigger than its own size is probably the reason its digs holes deep into the stuff it is digesting. Scraping and digging gets the job done faster than just employing a single approach.

The discovery was the unexpected result of very thorough imaging and analysis by Bryon Donohoe of NREL’s Biomass Surface Characterization Laboratory research team.

NREL researchers put CelA to the test and found that it produced more sugars than the most abundant cellulase in the leading commercial mixtures, Cel7A, when acting on Avicel, which is an industry standard to test cellulose degradation.

The team’s research paper on the finding, “Revealing Nature’s Cellulase Diversity: The Digestion Mechanism of Caldicellulosiruptor bescii CelA,” has been published in the journal Science.

It isn’t like the naturally occurring enzyme is the complete answer. For example, even though CelA contains naturally occurring endoglucanase, the researchers found that adding more endoglucanase greatly accelerated the rate at which it broke down sugars. CelA has its own beta-glucosidase activity, but NREL researchers checked to see how it would perform if more of the beta-glucosidase was added. That’s when the rate really accelerated.

Brunecky added, “You’d think that nature would already have evolved an optimal mix in a single enzyme. You expect to see maybe a 20% to 30% improvement when you add a beta glucosidase to a cocktail – not the doubling or tripling of the rate of conversion and the increase of rates by a factor of 10 that we got with CelA. Nobody expected the improvement to be this high.”

NREL Senior Scientist Yannick Bomble, who is the senior author of the Science paper explains, “The bacteria that secrete the promising CelA thrive in temperatures of 80°C to 90°C, close to boiling water. That’s an advantage because the early pre-treatment process requires temperatures greater than 100°C to remove unwanted materials. The next step requires cooling the temperature to the preferred range of the enzyme. In the case of CelA, the temperature doesn’t have to drop as much – another way to save money.”

“CelA is the most efficient single cellulase we’ve ever studied, by a large margin,” Bomble said. “It is an amazingly complex enzyme, combining two catalytic domains with three binding modules. The fact that it has two complementary catalytic domains working in concert most likely makes it such a good cellulose degrader.”

The discovery is getting support. In a letter of comment in the same issue of Science, Alex Berlin of biotechnology company Novozymes noted that CelA’s ability to operate at high temperatures “would be seen by many in the biomass biorefinery industry as an advantage” because it “dramatically reduces the chances of bacterial contamination” while lowering the viscosity of the mixtures.

The NREL researchers led by Michael Himmel aren’t resting on their laurels, or their patents. They’re examining the other enzymes secreted by the organism. They’re also using what they’ve learned from CelA to help improve cellulase enzymes that are more compatible with the enzyme formulations used today.

The next step toward using CelA at commercial scale is to express large amounts of the enzyme in existing production systems, and to boost enzyme yields from the native organism, C. bescii. Most of the large enzyme companies have the expertise to do that, so commercialization looks like a good bet.

If CelA proves reliable as it is scaled up to manufacturing levels, it could mean lower fuel prices in 2022, when the Energy Independence and Security Act mandates annual production of 36 billion gallons of fuel made from the non-food part of plants.

The discovery has immense potential implications. Lignocellulosic biomass is the most plentiful and sustainable resource on Earth, largely made up of plant residuals that would otherwise go unused and left to decay. Using this biomass as a source of alternative fuels can help offset the world’s dependence on fossil fuels and reduce greenhouse gas emissions.


3 Comments so far

  1. dmm on January 21, 2015 4:47 PM

    I am confused. That Science publication is from Dec2013 — more than a year ago. What has changed?

  2. Brian Westenhaus on January 21, 2015 9:38 PM

    Don’t know why it took NREL so long to announce the press release. Could be one or more of several reasons. No info on why, so the post went up. BW

  3. Al Fin on January 23, 2015 10:10 AM

    Low oil & gas prices may reduce the urgency of this type of research, but eventually it will be very important.

    High temperature nuclear reactors will eventually work hand in hand with advanced biofuels production, providing inexpensive process heat and power.

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