Characterization and Transcriptional Regulation of n-Alkane Hydroxylase Gene Cluster of Rhodococcus jostii RHA1

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

Complete genome sequence of natural rubber-degrading, gram-negative bacterium, Rhizobacter gummiphilus strain NS21T

Gram-negative natural rubber-degrader, Rhizobacter gummiphilus NS21T, which was isolated from soil in the botanical garden in Japan, is a newly proposed species of genus of Rhizobacter. It has been reported that the latA1 gene is involved in the natural rubber degradation in this strain. To gain novel insights into natural rubber degradation pathway, the complete genome sequence of this strain was determined. The genome of strain NS21T consists of 6,398,096 bp of circular chromosome (GenBank accession number CP015118.1) with G + C content of 69.72%. The genome contains 5687 protein-coding and 68 RNA genes. Among the predicted genes, 4810 genes were categorized as functional COGs. Homology search revealed that existence of latA1 homologous gene (latA2) in this genome. Quantitative reverse-transcription-PCR and deletion analyses indicated that natural rubber degradation of this strain requires latA2 as well as latA1. Keywords: Natural rubber, Rubber oxygenase, Gram-negative natural-rubber degrading bacteri

Characterization of Latex-Clearing Protein and Aldehyde Dehydrogenases Involved in the Utilization of poly(cis-1,4-isoprene) by Nocardia farcinica NBRC 15532

Microbial degradation of natural rubber and synthetic poly(cis-1,4-isoprene) is expected to become an alternative treatment system for waste from poly(cis-1,4-isoprene) products including scrap tires. Nocardia farcinica NBRC 15,532, a gram-positive rubber-degrading bacterium, can utilize poly(cis-1,4-isoprene) as the sole source of carbon and energy to produce oligo-isoprene metabolites containing aldehyde and keto end groups. A homology-based search of the genome revealed a gene encoding a latex-clearing protein (Lcp). Gene disruption analysis indicated that this gene is essential for the utilization of poly(cis-1,4-isoprene) in this strain. Further analysis of the genome sequence identified aldehyde dehydrogenase (ALDH) genes as potential candidates for oxidative degradation of oligo-isoprene aldehydes. Based on the enzymatic activity of the ALDH candidates, NF2_RS14000 and NF2_RS14385 may be involved in the degradation of oligo-isoprene aldehydes. Analysis of the reaction products revealed that these ALDHs oxidized tri- to penta-isoprene aldehydes, which were generated by the reaction of Lcp. Based on the inability of ALDH gene deletion mutants, we concluded that NF2_RS14000 is mainly involved in the utilization of poly(cis-1,4-isoprene) and the oxidative degradation of oligo-isoprene aldehydes in Nocardia farcinica NBRC 15,532