Building a better catalyst for bioremediation

Biotransformation of one substance into another using bacteria isn't something new. It has gone on as long as there has been life. But now, Los Alamos National Laboratory's Bioscience Division is trying to create a toolbox, of sorts, using bacteria to clean up industrial waste. For the national laboratories, that also means a way to clean up legacy waste from operations related to the nuclear weapons complex.

The idea seems simple enough: use an enzyme that will react with the contaminant, called a substrate, and convert it into something benign. The idea is appealing because the process could be used to treat contaminants otherwise too far underground to get at easily and economically. Jim Brainard and his team are concentrating on organic contaminants. The vast diversity of bacteria raises the prospect for the creation of a catalog of designer enzymes to use on waste. The process turns organics into more bacteria or something else such as carbon dioxide. Work on bioremediation of inorganic contaminants such as toxic metals, actinides and other man-made compounds also is in the early stages at the Lab.

"I think you can find an enzyme that will degrade almost any chemical in the environment," said Brainard. The team uses a combination of rational and evolutionary design approaches. With the rational approach, functional properties and structural features of different enzymes are compared and combined. Then, site-specific testing is done to see if results of the "marriage" of the enzymes accomplish desired goals. In the evolutionary design approach, the team makes large libraries of random mutations in proteins then picks out the enzymes that work well with a particular contaminant. The whole idea is to go from a good catalyst to a better catalyst.

So far, Brainard and his team have had success with a few well-known enzymes. Most are not the best catalysts for initiating contaminant transformation. So, some biological engineering has to take place to improve the efficiency of their reactions.

Many enzymes that transform toxic contaminants only work efficiently with high concentrations of contaminants. "Most of the known enzyme systems will not transform contaminants at low enough levels to catalyze transformations down to regulatory limits," Brainard noted. Brainard's colleagues are improving the interaction between an enzyme and contaminant so it will bind with the substrate at low concentrations. The benign product of an enzyme-contaminant reaction also might stop complete conversion of the contaminant altogether. "Product inhibition and binding of enzyme to substrate are both problems that need to be overcome if we are going to reach regulatory levels and be able to say we have 'cleaned' the site," says Brainard. Lab scientist Cliff Unkefer and his team have engineered enzymes that are no longer product inhibited. The designer enzymes will be carried to the site that need emediation by microbes.

Microbiologist Cheryl Kuske and her team are developing tools to better understand microbes in the environment (see "Microbial Diversity"). The majority of bacteria is something about which little is known because there are so many bacteria and most do not grow under laboratory conditions. Using a tree as an analogy, all animals make up just one small branch, plants are another and other microorganisms make up the rest. Microorganisms are the most diverse forms of life and more than 95 percent are still unknown. Another possible application for a library of "designer" enzymes is in the chemical industry. Biomaterials rather than petrochemicals could be used to develop feed stock. Glycerol for example, derived biologically, could be converted into a feed stock for many biochemicals. That raw material would be used to make polymers and plastics—all from the same enzyme process being developed to deal with organic chemical waste.

"That 's a money-maker rather than a cost. There are certainly more opportunities for research support if you have a target that potentially will make a chemical company money as opposed to a cost to clean up a site," Brainard said. "I would claim that research into better bioremediation tools is a national need. I think it's an important thing for us to do."