The goal of this Phase I SBIR research is to begin to elucidate the genetic and enzymatic mechanisms of methyl tert-butyl ether (MTBE) metabolism. MTBE has been identified as the second most common contaminant of urban ground waters in the United States, and it threatens the health of millions of Americans who may consume it through their drinking water. The results of this research will allow us to 1) better understand and predict the fate of this important groundwater contaminant in the environment; 2) better understand how tertiary carbon molecules like MTBE are metabolized in higher organisms; and 3) develop improved water treatment methods to reduce the risk of human exposure to MTBE through drinking water sources. During the six-month Phase I project we will clone, express, and sequence MTBE degradation genes from a recently isolated MTBE-degrading bacterium, strain ENV735. During follow on work we will evaluate the regulation of the MTBE-degrading genes to better understand how to improve in situ degradation of MTBE in groundwater, and to identify mechanisms for improving the performance of drinking water treatment systems to reduce human MTBE exposure. We also will evaluate the biochemistry of MTBE-degrading enzymes to help understand how human cells may metabolize small tertiary carbon molecules like MTBE