Gaseous and liquid alkanes can serve as bacterial growth substrates. Alkane monooxygenases initiate the metabolism of alkanes by catalyzing the oxidation of the alkanes to alcohols. While metabolism of methane and liquid alkanes (e.g., octane) is well studied, much less is known about the oxidation of short-chain, gaseous alkanes. The applicant's research has focused on BMOs as representatives of this understudied group of alkane utilizers and on the complete pathway of butane metabolism. Previous results with three butane-utilizing bacteria indicated a surprising level of diversity among the enzymes that harvest butane as well as those involved in the subsequent metabolism of the alcohols. The investigators now propose to focus on just one of these bacteria, Pseudomonas butanovora.
The specific aims of the project are: 1. Characterize the diiron containing BMO, which is similar to soluble MMO, but does not oxidize methane. 2. The applicants discovered two clusters of genes, each coding for a butanol dehydrogenase and a putative butyraldehyde dehydrogenase, which use parallel pathways of electron transport. The applicant now proposes to identify the components of these parallel pathways. 3. Previous studies indicated independent regulation of the production of BMO, butanol dehydrogenases and butyraldehyde dehydrogenases by butane and intermediates in the pathway. The investigators will examine the molecular basis of the regulation of induction of the butane pathway in P. butanovora. The proposed experiments may provide unique insights to the basis of substrate specificity in the broad class of diiron monooxygenases that oxidize nonpolar molecules. Studies of parallel pathways will be relevant to other metabolic pathways where toxic metabolites are necessarily produced as part of the metabolism of a growth supporting substrate.
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