Hydrogen Autotrophy as a Transferable Genetic Character of Nocardia opaca 1b

Reh, Michael; Schlegel, H. G.

doi:10.1099/00221287-126-2-327

Cited by 13 publications

(8 citation statements)

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“…element (Reh & Schlegel, 1981). This assumption was confirmed by determining the specific activities of relevant enzymes in the donors and autotrophic transconjugants (Ecker e l al., 1986).…”

Section: Introductionmentioning

confidence: 59%

The giant linear plasmid pHG207 from Rhodococcus sp. encoding hydrogen autotrophy: characterization of the plasmid and its termini

Kalkus

Dörrie

Fischer

et al. 1993

Journal of General Microbiology

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As described previously, in Rhodococcus sp. (formerly Nocardia opaca) strains MRll and MR22, the ability to grow as an aerobic hydrogen bacterium (the Aut character) is located on giant conjugative linear plasmids -on pHG201 (270 kb) in strain MRll and on pHG205 (280 kb) in strain MR22. In an autotrophic transconjugant originating from MR22 a smaller plasmid, pHG207 (225 kb), was detected and shown to be a recombination product of the wild-type plasmids pHG204 and pHG205. A donor carrying pHG207 as the sole plasmid transferred the Aut marker at a 1000-fold frequency compared to the wild-type plasmid pHG205. Analysis of the plasmid ends revealed that plasmid pHG207 carries proteins at both ends; the proteins are linked to the 5' ends of the strands. The cloned end fragments of about 2 kb were sequenced and found to contain highly homologous sequences within the terminal 583 bp (left end part) and 560 bp (right end part). Several potential reading frames were detected, but database searching gave no indication about possibIe functions.

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Section: Introductionmentioning

confidence: 59%

The giant linear plasmid pHG207 from Rhodococcus sp. encoding hydrogen autotrophy: characterization of the plasmid and its termini

Kalkus

Dörrie

Fischer

et al. 1993

Journal of General Microbiology

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“…3). Instead, the proteins of A. eutrophus, Rhodobacter sphaeroides, and X. flavus, which belong to the ␣ and ␤ subdivisions of the proteobacteria, have the protein from (3,7,33,41). Plasmids harboring cbb genes which can be transferred via conjugation to other bacteria are frequently encountered in gram-negative bacteria (12,16,38,52).…”

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confidence: 99%

Primary structure and phylogeny of the Calvin cycle enzymes transketolase and fructosebisphosphate aldolase of Xanthobacter flavus

et al. 1996

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Xanthobacter flavus grows autotrophically by using the Calvin cycle for the fixation of CO 2 . Only 2 of the 11 enzymes of the Calvin cycle are characteristic for this pathway; the others are also present during heterotrophic growth. The key enzymes of the Calvin cycle, phosphoribulokinase and ribulosebisphosphate carboxylase, are encoded within the cbb operon, which is transcribed only during autotrophic growth. Two additional genes are located within this operon: cbbX, encoding a protein with unknown function, and cbbF, encoding fructosebisphosphatase (27,29). The transcription of the cbb operon is positively regulated by CbbR, a LysR-type transcriptional regulator, which binds to two sites in the cbb promoter (47).During autotrophic growth, X. flavus uses two fructosebisphosphatase enzymes with distinct properties. The inducible enzyme encoded by cbbF has a high level of sedoheptulosebisphosphatase activity and is stimulated by ATP. The second constitutive fructosebisphosphatase has a low level of sedoheptulosebisphosphatase activity and is not stimulated by ATP (48). In contrast to the fructosebisphosphatase isoenzyme pair, only one phoshoglycerate kinase gene, which is not encoded within the cbb operon, is employed by X. flavus. The pgk gene is constitutively expressed, but the expression level is higher during autotrophic growth than during heterotrophic growth (26).Little is known about the transketolase (EC 2.2.1.1) and fructosebisphosphate aldolase (FBP aldolase; EC 4.1.2.13) enzymes of X. flavus. Like fructosebisphosphatase and phosphoglycerate kinase, these enzymes are involved in both heterotrophic metabolism and the fixation of CO 2 via the Calvin cycle. Two unrelated mechanistically distinct types of FBP aldolase enzymes are encountered in bacteria, archaea, and eukarya (24). Class I FBP aldolases form a covalent Schiff base between the substrate and the ␣-amino group of a lysine residue during catalysis, whereas the class II enzymes depend on a divalent cation as the electrophile in the catalytic cycle (24).X. flavus (Table 1) was grown heterotrophically on succinate (10 mM) or gluconate (10 mM) and autotrophically on methanol (0.5% [vol/vol]) at 30ЊC as described previously (22,28). The activities of transketolase and FBP aldolase were determined, according to published methods (15, 49), in cell extracts which were prepared by using a French pressure cell as described previously (27). The activity of transketolase was increased sixfold following autotrophic growth on methanol compared with that of heterotrophically grown cells. In sharp contrast, the activity of FBP aldolase (without Fe 2ϩ ) was the same for both heterotrophic and autotrophic growth. Because the activity of class II FBP aldolase is dependent on Fe 2ϩ as the electrophile (24), FeSO 4 (700 M) was included in the reaction assay. Surprisingly, Fe 2ϩ did not affect the FBP aldolase activity in the cell extracts of heterotrophically grown cells but stimulated the activity of FBP aldolase in cell extracts of autotrophically grown cells ...

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“…R. opacus strain MR11, originally isolated by D. Siebert, is the type strain (Aggag & Schlegel, 1973;Klatte et al, 1994). The derivative strains MR2246 and MR1463 were described (Sensfuss et al, 1986;Reh & Schlegel, 1981) and studied in this laboratory.Escherichia coli BMH71-18, plasmid pMc5-8 and A. eutrophus HF14 were provided by W. Kramer, H.-J. Fritz and B. Friedrich, respectively.…”

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Genes encoding the NAD-reducing hydrogenase ofRhodococcus opacus MR11

et al. 1997

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The dissociation of the soluble NAD-reducing hydrogenase of Rhodococcus opacus M R l l into two dimeric proteins with different catalytic activities and cofactor composition is unique among the NAD-reducing hydrogenases studied so far. The genes of the soluble hydrogenase were localized on a 7-4 kbp Asnl fragment of the linear plasmid pHG2Ol via heterologous hybridization. Analysis of the nucleotide sequence of this fragment revealed the seven open reading frames ORF1, hoxF, -U, -Y, -H, -Wand ORF7. The six latter ORFs belong to the gene cluster of the soluble hydrogenase. Their gene products are highly homologous to those of the NAD-reducing enzyme of Alcaligenes eutrophus H16. The genes hoxF, -U, -Y and -H encode the subunits a, 7, 6 and B, respectively. The gene hoxW encodes a putative protease, which may be essential for C-terminal processing of the / 3 subunit. Finally, ORF7 encodes a protein which has similarities to CAMP-and cGMP-binding protein kinases, but its function is not known. ORFI, which lies upstream of the hydrogenase gene cluster, encodes a putative transposase found in IS elements of other bacteria. Northern hybridizations and primer extensions using total RNA of autotrophically and heterotrophically grown cells of R. opacus M R l l indicated that the hydrogenase genes are under control of a like promoter located at the right end of ORFI and are even transcribed under heterotrophic conditions a t a low level. Furthermore, this promoter was shown to be active in the recombinant Escherichia coli strain LHYl harbouring the 7.4 kbp Asnl fragment, resulting in overexpression of the hydrogenase genes. Although all four subunits of the soluble hydrogenase were shown via Western immunoblots to be synthesized in E. coli, no active enzyme was detectable.Keywords : Rhodococcus opacus MR11, NAD-reducing hydrogenase, box genes INTRODUCTIONRhodococcus opacus strain M R l l (formerly Nocardia opaca 1 b) is a Gram-positive, facultative chemolithoautotrophic bacterium which can grow on carbon dioxide and gaseous H, as the sole carbon and energy sources. Physiologically, it belongs to the knallgas bacteria, a group which is composed of phylogenetically diverse organisms. For the activation of H,, all knallgas bacteria contain hydrogenases, which are of two basic types : the membrane-bound hydrogenases (MBHs) , which are coupled to the electron transfer chain and are The GenBank accession number for the nucleotide sequence reported in this paper is U70364.not capable of reducing NAD, and the soluble hydrogenases (SHs), which are localized in the cytoplasm and catalyse the transfer of electrons directly to NAD (Aragno & Schlegel, 1992;Schneider et al., 1984a;Zaborosch et al., 1989; Schink & Schlegel, 1979). The MBHs belong to the more common group of NiFehydrogenases consisting of one large and one small subunit, whereas the SHs belong to a family of less abundant multimeric NiFe-hydrogenases (Friedrich & Schwartz, 1993 ;Wu & Mandrand, 1993). The majority of the knallgas bacteria contain only the MBH, but a few, ...

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Hydrogen Autotrophy as a Transferable Genetic Character of Nocardia opaca 1b

Cited by 13 publications

References 18 publications

The giant linear plasmid pHG207 from Rhodococcus sp. encoding hydrogen autotrophy: characterization of the plasmid and its termini

The giant linear plasmid pHG207 from Rhodococcus sp. encoding hydrogen autotrophy: characterization of the plasmid and its termini

Primary structure and phylogeny of the Calvin cycle enzymes transketolase and fructosebisphosphate aldolase of Xanthobacter flavus

Genes encoding the NAD-reducing hydrogenase ofRhodococcus opacus MR11

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