6533b85cfe1ef96bd12bc975

RESEARCH PRODUCT

Involvement of an Alkane Hydroxylase System of Gordonia sp. Strain SoCg in Degradation of Solid n-Alkanes▿

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ABSTRACT Enzymes involved in oxidation of long-chain n -alkanes are still not well known, especially those in Gram-positive bacteria. This work describes the alkane degradation system of the n -alkane degrader actinobacterium Gordonia sp. strain SoCg, which is able to grow on n -alkanes from dodecane (C 12 ) to hexatriacontane (C 36 ) as the sole C source. SoCg harbors in its chromosome a single alk locus carrying six open reading frames (ORFs), which shows 78 to 79% identity with the alkane hydroxylase (AH)-encoding systems of other alkane-degrading actinobacteria. Quantitative reverse transcription-PCR showed that the genes encoding AlkB (alkane 1-monooxygenase), RubA3 (rubredoxin), RubA4 (rubredoxin), and RubB (rubredoxin reductase) were induced by both n -hexadecane and n -triacontane, which were chosen as representative long-chain liquid and solid n -alkane molecules, respectively. Biotransformation of n -hexadecane into the corresponding 1-hexadecanol was detected by solid-phase microextraction coupled with gas chromatography-mass spectrometry (SPME/GC-MS) analysis. The Gordonia SoCg alkB was heterologously expressed in Escherichia coli BL21 and in Streptomyces coelicolor M145, and both hosts acquired the ability to transform n -hexadecane into 1-hexadecanol, but the corresponding long-chain alcohol was never detected on n -triacontane. However, the recombinant S. coelicolor M145-AH, expressing the Gordonia alkB gene, was able to grow on n -triacontane as the sole C source. A SoCg alkB disruption mutant that is completely unable to grow on n -triacontane was obtained, demonstrating the role of an AlkB-type AH system in degradation of solid n -alkanes.