| Summary: | Gram-positive actinomycete <i>Rhodococcus jostii</i> RHA1 is able to grow on C10 to C19 <i>n</i>-alkanes as a sole source of carbon and energy. To clarify, the <i>n</i>-alkane utilization pathway—a cluster of 5 genes (<i>alkBrubA1A2BalkU</i>) which appeared to be involved in <i>n</i>-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 <i>n</i>-alkane. Inactivation of <i>alkB</i> led to the absence of the ability to utilize <i>n</i>-undecane. The <i>alkB</i> mutation resulted in reduction of growth rates on C10 and C12 <i>n</i>-alkanes; however, growths on C13 to C19 <i>n</i>-alkanes were not affected by this mutation. These results suggested that <i>alkB</i> was essential for the utilization of C10 to C12 <i>n</i>-alkanes. Inactivation of <i>alkU</i> showed the constitutive expression of <i>alkB</i>. Purified AlkU is able to bind to the putative promoter region of <i>alkB</i>, suggesting that AlkU played a role in repression of the transcription of <i>alk</i> operon. The results of this study indicated that <i>alkB</i> was involved in the medium-chain <i>n</i>-alkanes degradation of strain RHA1 and the transcription of <i>alk</i> operon was negatively regulated by <i>alkU</i>-encoded regulator. This report is important to understand the <i>n</i>-alkane degradation pathway of <i>R. jostii</i>, including the transcriptional regulation of <i>alk</i> gene cluster.
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