Abstract
Gram-positive actinomycete Rhodococcus jostii RHA1 is able to grow on C10 to C19 n-alkanes as a sole source of carbon and energy. To clarify, the n-alkane utilization pathway—a cluster of 5 genes (alkBrubA1A2BalkU) which appeared to be involved in n-alkane degradation—was identified and the transcriptional regulation of these genes was characterized. Reverse transcription-PCR analyses revealed that these genes constituted an operon and were transcribed in the presence of n-alkane. Inactivation of alkB led to the absence of the ability to utilize n-undecane. The alkB mutation resulted in reduction of growth rates on C10 and C12 n-alkanes; however, growths on C13 to C19 n-alkanes were not affected by this mutation. These results suggested that alkB was essential for the utilization of C10 to C12 n-alkanes. Inactivation of alkU showed the constitutive expression of alkB. Purified AlkU is able to bind to the putative promoter region of alkB, suggesting that AlkU played a role in repression of the transcription of alk operon. The results of this study indicated that alkB was involved in the medium-chain n-alkanes degradation of strain RHA1 and the transcription of alk operon was negatively regulated by alkU-encoded regulator. This report is important to understand the n-alkane degradation pathway of R. jostii, including the transcriptional regulation of alk gene cluster.
Highlights
Alkanes constitute up to 50% of crude oil and are commonly found in oil-contaminated environments [1]
In the case of Acinetobacter baylyi ADP1, a three-component alkane hydroxylase containing alkane 1-monooxygenase encoded by alkM, a rubredoxin encoded by rubA, and a rubredoxin reductase encoded by rubB, which is similar to the GPo1 enzyme, catalyzes terminal
We found that RHA1 utilized C10 to C19 n-alkanes, and characterized the operon structure and transcriptional regulation of n-alkane degradation genes in RHA1
Summary
Alkanes constitute up to 50% of crude oil and are commonly found in oil-contaminated environments [1] Various microorganisms, both aerobic and anaerobic, utilize alkanes as a sole carbon and energy source [2,3]. The alkane-hydroxylation mechanism of Pseudomonas putida GPo1 has been characterized in detail, and consists of three subunits: an integral-membrane non-heme di-iron monooxygenase encoded by alkB; a rubredoxin encoded by duplicated genes, alkF and alkG; and a rubredoxin reductase encoded by alkT [8]. These genes are distributed in two different loci of the OCT plasmid in the strain GPo1 [8]. In the case of Acinetobacter baylyi ADP1, a three-component alkane hydroxylase containing alkane 1-monooxygenase encoded by alkM, a rubredoxin encoded by rubA, and a rubredoxin reductase encoded by rubB, which is similar to the GPo1 enzyme, catalyzes terminal
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