Elucidation of multiple alkane hydroxylase systems in biodegradation of crude oil n ‐alkane pollution by Pseudomonas aeruginosa DN1

The purpose of this study was to elucidate the characteristics of multiple alkane hydroxylase systems in Pseudomonas aeruginosa DN1, including two homologues of AlkB (AlkB1 and AlkB2), a CYP153 homologue (P450), and two homologues of Alm‐like (AlmA1 and AlmA2). DN1 was capable of utilizing diverse n‐alkanes with chain lengths from 8 to 40 C atoms as the sole carbon source, and displayed high degradation efficiency (>85%) of crude oil and a majority of n‐alkanes using gas chromatography method. RT‐qPCR analysis showed that the five enzyme genes could be induced by n‐alkanes ranging from medium‐chain length to long‐chain length which indicated the dissimilarity of expression between those genes when grown on different n‐alkanes. Notably, the expression of alkB2 gene was upregulated in the presence of all of the tested n‐alkanes, particularly responded to long‐chain n‐alkanes like C20 and C32. Meanwhile, long‐chain n‐alkanes (C20‐C36) significantly elevated cyp153 expression level, and the expression of two almA genes was only upregulated in the presence of n‐alkanes with chain lengths of 20C's and longer. Furthermore, the disruption of those genes demonstrated that AlkB2 appeared to play a key role in the biodegradation of substrates of a broad‐chain length ranges, besides other alkane hydroxylase systems ensured the utilization of n‐alkanes with chain lengths of from 20 to 40 C atoms. The five functional alkane hydroxylase genes make DN1 an attractive option for its versatile alkane degradation, which is primarily dependent on the expression of alkB2. Our findings suggest that P. aeruginosa DN1 is a predominately potential long‐chain n‐alkane‐degrading bacterium with multiple alkane hydroxylase systems in crude oil‐contaminated environment.