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NMR Analysis of the Anaerobic Degradation Pathway of Toluene in Azoarcus tolulyticus
EPA Grant Number: U915388Title: NMR Analysis of the Anaerobic Degradation Pathway of Toluene in Azoarcus tolulyticus
Investigators: Griffin, Benjamin M.
Institution: Michigan State University
EPA Project Officer: Just, Theodore J.
Project Period: January 1, 1998 through January 1, 2001
Project Amount: $88,633
RFA: STAR Graduate Fellowships (1998)
Research Category: Fellowship - Microbiology , Academic Fellowships , Biology/Life Sciences
Description:
Objective:The objective of this research project is to perform structural analyses of intermediates in anaerobic toluene degradation by Azoarcus tolulyticus to identify metabolites and explore key enzyme mechanisms in this pathway. The results of this project will expand our knowledge of the fate of toluene in anaerobic environments, and will aid in the design of remediation strategies.
Approach:One major challenge in the study of microbial degradation of pollutants is the detection of metabolites, which may be present in low concentrations or unstable during sample preparation and analysis procedures. To avoid these problems during the study of toluene degradation, I am using nuclear magnetic resonance (NMR) spectroscopy as my primary analytical technique. NMR has several advantages because it is noninvasive, nondestructive, and requires no purification or extraction steps before analysis. This allows for the identification of multiple pathway intermediates in a single assay on intact samples. I am studying the anaerobic degradation of toluene in A. tolulyticus strain Tol-4. Strain Tol-4 is the designated type strain of the newly discovered genus Azoarcus, which recent investigations have demonstrated to be quite common in nature. Specifically, I am tracking the transformation of 13C-, 2H-, and 19F-labeled toluenes in resting cells and cell-free extracts of strain Tol-4. A suite of structural analogs, including xylenes and fluorotoluenes, will be used to selectively block portions of the pathway to increase the accumulation of intermediates. Because NMR gives detailed structural information, analysis of labeled products will provide insight into both the identity of intermediates and, in certain cases, the reaction mechanism.
Supplemental Keywords:fellowship, pollutants, microbial degradation, toluene, anaerobic degradation, remediation strategies, nuclear magnetic resonance, NMR, Azoarcus tolulyticus, enzyme mechanisms.