Biochemistry of Two FADH2-Utilizing Trichlorophenol 4-Monooxygenases

Washington State University, Pullman WA

Investigators

Abstract

Polychlorophenols are a major group of pollutants, which have been introduced into the environment over the last century through their applications in consumer products, wood industry, and agriculture. Due to special uses for compounds with particular substitutions and due to lack of specificity in some of the manufacturing processes, many polychlorophenol isomers exist. Often the importance of a halide substitution is that it increases the persistence of a compound by making it relatively recalcitrant to microbial degradation. Too much success in making a persistent compound has led to its accumulation in the environment. Efforts are underway to identify and develop microorganisms that can degrade some of the more problematic polychlorophenols. Research from the PI's group has significantly contributed to the complete elucidation of degradation pathways for three important polychlorophenols: 2,4,6-trichlorophenol (2,4,6-TCP), 2,4,5-trichlorophenol (2,4,5-TCP), and pentachlorophenol. A novel group of FADH2-utilizing monooxygenases that oxidize polychlorophenols has been discovered. Two closely related FADH2-utilizing monooxygenases, TcpA and TftD, initiate the degradation of 2,4,6-TCP and 2,4,5-TCP, respectively. Recent data suggests that TcpA catalyzes both oxidative and hydrolytic reactions for two dechlorination steps, whereas TftD sequentially dechlorinates 2,4,5-TCP by two oxidation reactions. The first research objective is to demonstrate the proposed dechlorination mechanisms. The second objective is to localize the amino acid residues responsible for the altered catalysis of the highly homologous TftD and TcpA by a tailored domain swapping and site-directed mutagenesis strategy. The results would lead to a more complete description of the way, in which enzymes are recruited by mutation and selection to degrade novel compounds. The proposed research aims to provide scientific guidance for applying FADH2-utilizing monooxygenases in bioremediation of environmental pollutants. The project investigates how microorganisms are evolving to degrade various isomers of polychlorophenols, a group of highly toxic pollutants. The project also studies the reaction mechanisms employed by microorganisms to degrade these compounds. The research will have a broad impact on society because the results will provide scientific guidance for the remediation of polychlorophenols. The research will also have a direct impact on biochemistry, microbiology, and evolution theories. Through this research, a graduate student will be trained to perform independent research in biochemistry and microbiology. Several undergraduate students will work on some aspects of the project to gain research experience. These educational efforts will prepare students to meet new challenges in biological and environmental sciences.