Metabolic and stress responses of Acinetobacter oleivorans DR1 during long-chain alkane degradation

Acinetobacter oleivorans DR1 can utilize C₁₂–C₃₀ alkanes as a sole carbon source but not short-chain alkanes (C₆, C₁₀). Two copies of each alkB-, almA- and ladA-type alkane hydroxylase (AH) are present in the genome of DR1 cells. Expression and mutational analyses of AHs showed that alkB1 and alkB2 are the major AH-encoding genes under C₁₂–C₃₀, and the roles of other almA- and ladA genes are negligible. Our data suggested that AlkB1 is responsible for long-chain alkane utilization (C₂₄–C₂₆), and AlkB2 is important for medium-chain alkane (C₁₂–C₁₆) metabolism. Phylogenetic analyses revealed large incongruities between phylogenies of 16S rRNA and each AH gene, which implies that A. oleivorans DR1 has acquired multiple alkane hydroxylases through horizontal gene transfer. Transcriptomic and qRT-PCR analyses suggested that genes participating in the synthesis of siderophore, trehalose and poly 3-hydroxybutyrate (PHB) were expressed at much higher levels when cells used C₃₀ than when used succinate as a carbon source. The following biochemical assays supported our gene expression analyses: (i) quantification of siderophore, (ii) measurement of trehalose and (iii) observation of PHB storage. Interestingly, highly induced both ackA gene encoding an acetate kinase A and pta gene encoding a phosphotransacetylase suggested unusual ATP synthesis during C₃₀ alkane degradation, which was demonstrated by ATP measurement using the ΔackA mutant. Impaired growth of the ΔaceA mutant indicated that the glyoxylate shunt pathway is important when C₃₀ alkane is utilized. Our data provide insight into long-chain alkane degradation in soil microorganisms.