Mining for new enzymes

to create bioplastics

Published December 01 2017

This extreme heat-resistant polymer was synthesized by Tatsuo Kaneko of the Japan Advanced Institute of Science and Technology from an aromatic compound. The nitrating-P450 found by the Ohnishi group may be useful in the microbial production of a monomer for such a super-engineering plastic.courtesy of Tatsuo Kaneko

Plastics are a ubiquitous part of our daily lives. Most plastics that we use are the product of petroleum or natural gas. Not only are plastics made of petroleum or natural gas, but they also require additional fossil fuels to produce. Further, our infatuation with plastics is increasing; more plastic was produced globally in the past 13 years than in the previous 50.

Yasuo Ohnishi and colleagues from the University of Tokyo and Ishikawa Prefectural University aim to use bacteria to create starting materials for production of super-engineering plastics (those with better mechanical and thermal properties than more widely used commodity plastics), decreasing a reliance on fossil fuels. Toward this goal, in a recent paper in the Journal of Biological Chemistry, Ohnishi and colleagues write that they have identified a new enzyme that can add a nitro group to the benzene ring of tyrosine to produce 3-nitrotyrosine. A nitro group can easily be reduced to an amino group, which is a useful functional group for polymer production. This finding has the potential to be a new avenue toward using bacteria to create useful compounds in bioplastic development as well as helping to design metabolic pathways in the field of synthetic biology.

Benzene is a ring of six carbon atoms that can be used as a starting material to make materials such as polystyrene, nylon and adhesives. The Ohnishi lab was awarded a grant from the Japan Science and Technology Agency to create new compounds that can be used for bioplastics production or in the field of synthetic biology. As Ohnishi described it, “In the microbial production of useful compounds, the microbial cell is a ‘factory’ and enzymes are ‘machines.’ So, we needed a new machine to design a new ‘assembly line’ to produce aromatic compounds that can be used as a monomer of bioplastics; the new machine that we needed is an aromatic compound-nitrating enzyme.”

To find this machine, the Ohnishi group initiated a search of a chemical compound database to identify nitrated benzene compounds. This search identified rufomycin, a small circular molecule containing seven amino acids that was first identified as an anti-tuberculosis drug. Most relevant for the studies by the Ohnishi group, rufomycin contains a nitrated tyrosine residue. Thus, the Ohnishi group was able to identify the nitrating enzyme by isolating the genes that make rufomycin, which enabled the discovery of a new protein, RufO, a tyrosine 3-nitration enzyme.

The lab’s excitement at finding the first enzyme to nitrate tyrosine residues is tempered slightly by the observation that the enzyme activity is not very high. To integrate RufO into a useful enzyme for bioplastic production, it needs to have higher activity. To this end, the Ohnishi lab hopes to use two approaches to improve tyrosine nitration. First, now that a protein has been identified that performs this function, searches of homologous genes may identify proteins with related functionality. Second, structural analysis and mutagenesis studies may provide a pathway to generate new proteins with enhanced functionality. If either of these approaches is successful, we will be one step closer to decreasing our reliance on petroleum and natural gas for making plastics.

Robert E. Dempski Robert E. Dempski is an associate professor in the department of chemistry and biochemistry at the Worcester Polytechnic Institute.