Project description:Rhodococcus rhodochrous ATCC 17895 possesses an array of mono- and dioxygenases, as well as hydratases, which makes it an interesting organism for biocatalysis. R. rhodochrous is a Gram-positive aerobic bacterium with a rod-like morphology. Here we describe the features of this organism, together with the complete genome sequence and annotation. The 6,869,887 bp long genome contains 6,609 protein-coding genes and 53 RNA genes. Based on small subunit rRNA analysis, the strain is more likely to be a strain of Rhodococcus erythropolis rather than Rhodococcus rhodochrous.
Project description:The bacterial isolates of genus Rhodococcus are best known for their significant biodegradation abilities. Here, we report the data related to draft genome sequencing of Rhodococcus rhodochrous strain SPC17 isolated from sediments of Lonar Lake. The de novo assembly of 1598096 Illumina's paired-end sequencing reads resulted in 51 contigs for an overall genome assembly size of 4.98Mb. A total of 4546 genes were predicted using the National Center for Biotechnology Information- Prokaryotic Genome Annotation Pipeline (NCBI-PGAP). RAST server-based annotation of the Rhodococcus strain SPC17 genome resulted in a total of 295 subsystems with 25% subsystem coverage. The data on the draft genome shotgun project are accessible at NCBI-GenBank under the accession number WUUR00000000. Our data resource will facilitate further molecular and genomic studies of diverse hydrocarbon catabolizing genes present in Rhodococcus rhodochrous strain SPC17.
Project description:Rhodococcus rhodochrous PY11 (DSM 101666) is able to use 2-hydroxypyridine as a sole source of carbon and energy. By investigating a gene cluster (hpo) from this bacterium, we were able to reconstruct the catabolic pathway of 2-hydroxypyridine degradation. Here, we report that in Rhodococcus rhodochrous PY11, the initial hydroxylation of 2-hydroxypyridine is catalyzed by a four-component dioxygenase (HpoBCDF). A product of the dioxygenase reaction (3,6-dihydroxy-1,2,3,6-tetrahydropyridin-2-one) is further oxidized by HpoE to 2,3,6-trihydroxypyridine, which spontaneously forms a blue pigment. In addition, we show that the subsequent 2,3,6-trihydroxypyridine ring opening is catalyzed by the hypothetical cyclase HpoH. The final products of 2-hydroxypyridine degradation in Rhodococcus rhodochrous PY11 are ammonium ion and ?-ketoglutarate.
Project description:Rhodococcus rhodochrous is a bacterial species with applications in biocatalysis and bioremediation. Here, we report the draft genome sequence of strain G38GP, isolated from the gut of the cockroach Gromphadorhina portentosa. The genome consists of 76 contigs, with a total length of 6,256,198 bp and a GC content of 67.82%.
Project description:The 1.4-kb downstream region from a nitrilase gene (nitA) of an actinomycete Rhodococcus rhodochrous J1, which is industrially in use, was found to be required for the isovaleronitrile-dependent induction of nitrilase synthesis in experiments using a Rhodococcus-Escherichia coli shuttle vector pK4 in a Rhodococcus strain. Sequence analysis of the 1.4-kb region revealed the existence of an open reading frame (nitR) of 957 bp, which would encode a protein with a molecular mass of 35,100. Deletion of the central and 3'-terminal portion of nitR resulted in the complete loss of nitrilase activity, demonstrating that nitR codes for a transcriptional positive regulator in nitA expression. The deduced amino acid sequence of nitR showed similarity to a positive regulator family including XylS from Pseudomonas putida and AraC from E. coli. By Northern blot analysis, the 1.4-kb transcripts for nitA were detected in R. rhodochrous J1 cells cultured in the presence of isovaleronitrile, but not those cultured in the absence of isovaleronitrile. The transcriptional start site for nitA was mapped to a C residue located 26 bp upstream of its translational start site. Deletion analysis to define the nitA promoter region suggested the possible participation of an inverted repeat sequence, centered on base pair -52, in induction of nitA transcription.
Project description:Nitrile hydratase (NHase) from Rhodococcus rhodochrous J1 is widely used for industrial production of acrylamide and nicotinamide. However, the two types of NHases (L-NHase and H-NHase) from R. rhodochrous J1 were only slightly expressed in E. coli by routine methods, which limits the comprehensive and systematic characterization of the enzyme properties. We successfully expressed the two types of recombinant NHases in E. coli by codon-optimization, engineering of Ribosome Binding Site (RBS) and spacer sequences. The specific activity of the purified L-NHase and H-NHase were 400 U/mg and 234 U/mg, respectively. The molecular mass of L-NHase and H-NHase was identified to be 94 kDa and 504 kDa, respectively, indicating that the quaternary structure of the two types of NHases was the same as those in R. rhodochrous J1. H-NHase exhibited higher substrate and product tolerance than L-NHase. Moreover, higher activity and shorter culture time were achieved in recombinant E. coli, and the whole cell catalyst of recombinant E. coli harboring H-NHase has equivalent efficiency in tolerance to the high-concentration product relative to that in R. rhodochrous J1. These results indicate that biotransformation of nitrile by R. rhodochrous J1 represents a potential alternative to NHase-producing E. coli.
Project description:We recently reported that the overexpression of GroEL2 played an important role in increasing the alkane tolerance of Rhodococcus erythropolis PR4. In the present study, we examined the effects of the introduction of groEL2 on the alkane tolerance of other Rhodococcus strains. The introduction of groEL2 into Rhodococcus strains led to increased alkane tolerance. The translocation of R. rhodochrous ATCC12674 cells to and survival in the n-octane (C8) phase in two phase culture were significantly enhanced by the introduction of groEL2 derived from strain PR4, suggesting that engineering cells to overexpress GroEL2 represents an effective strategy for enhancing organic solvent tolerance in Rhodococcus.