Researchers at the Indian Institute of Science’s (IISc) Department of Inorganic and Physical Chemistry (IPC) created an enzymatic platform that can effectively convert naturally occurring, cheap fatty acids into valuable hydrocarbons known as 1-alkenes, which are promising biofuels. The study has been published in Science Advances.
Engineered whole cell biocatalyst for efficient 1-alkene production. Image Credit: Tabish Iqbal
Scientists are looking more closely at sustainable fuel pathways involving molecules known as hydrocarbons due to the limited supply and harmful effects of fossil fuels. When combined and utilized with currently available fuels and infrastructure, they exhibit significant promise as "drop-in" biofuels.
These hydrocarbons can potentially be synthesized on a large scale using microorganism "factories." As a result, enzymes that facilitate the mass production of these hydrocarbons are in high demand. Hydrocarbons are also extensively used in the polymer, detergent, and lubricant industries.
The IISc group previously isolated and studied an enzyme called UndB attached to the membranes of living cells, particularly some bacteria. It has the fastest rate of conversion of fatty acids to 1-alkenes currently achievable. However, the team discovered that the procedure was ineffective because the enzyme would inactivate after a short number of cycles. After conducting more research, they discovered that UndB was being inhibited by H2O2, a byproduct of the reaction process.
The catalase degrades the H2O2 that is produced.
Tabish Iqbal, Study First Author and Ph.D. Student, Department of Inorganic and Physical Chemistry, Indian Institute of Science
He says that adding catalase enhanced the enzyme's activity 19-fold, increasing from 14 to 265 turnovers (where turnover indicates the number of active cycles an enzyme completes before becoming inactivated).
Motivated by this discovery, the group used plasmid carriers to introduce a fused genetic code into E. coli bacteria, generating an artificial fusion protein combining UndB and catalase. These E. coli would then function as a "whole cell biocatalyst," converting fatty acids and churning out alkenes under the correct circumstances.
However, there were some difficulties. Since UndB is a membrane protein, working with it can be difficult. In excess of a specific concentration, it may be harmful to bacterial cells. Maintaining the proper conditions for studying membrane proteins like UndB is challenging because they are not soluble in water.
The researchers investigated the impact of several "redox partner" proteins that assist in moving electrons from fatty acids to alkenes to increase the effectiveness of their chimeric protein. The most effective proteins for providing electrons were discovered to be ferredoxin and ferredoxin reductase, as well as nicotinamide adenine dinucleotide phosphate (NADPH). When these were added to the genetically altered E. coli and fed fatty acids, the conversion efficiency increased to 95%.
A vital advantage of this biocatalyst is its specificity; UndB produces only pure 1-alkene without any unwanted side products, says Debasis Das, Assistant Professor at IPC and corresponding author.
1-alkenes can directly be used as biofuels.
Debasis Das, Assistant Professor and Study Corresponding Author, Department of Inorganic and Physical Chemistry, Indian Institute of Science
The group discovered that various fatty acids with different carbon chain configurations could be converted to 1-alkene by their biocatalyst. The group also demonstrated that the biocatalyst can produce styrene, a crucial product in the chemical and polymer industries.
The group has submitted a patent application for its whole cell biocatalyst and engineered protein. It is also searching for industry partners to expand the platform and enable mass production.
Our platform can be efficiently used to generate a large number of 1-alkenes that are valuable in biotechnology and polymer industries.
Debasis Das, Assistant Professor and Study Corresponding Author, Department of Inorganic and Physical Chemistry, Indian Institute of Science
Journal Reference:
Iqbal, T., et al. (2024) A chimeric membrane enzyme and an engineered whole-cell biocatalyst for efficient 1-alkene production. Science Advances. doi.org/10.1126/sciadv.adl2492