The hydrogenase gene cluster ofRhizobium leguminosarum bv.viciae contains an additional gene (hypX), which encodes a protein with sequence similarity to the N10-formyltetrahydrofolate-dependent enzyme family and is required for nickel-dependent hydrogenase processing and activity

Rey, Luís; Fernández, Domingo; Brito, Belén; Hernando, Yolanda; Palacios, José Luis Calvo; Imperial, Juan; Ruiz-Argüeso, Tomás

doi:10.1007/bf02173769

Cited by 32 publications

(40 citation statements)

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“…This construct complemented hydrogenase activity to wild-type levels regardless of the orientation of the insert relative to that of the lac promoter on the vector, indicating that the promoter region for hoxX is located within 800 bp upstream of the start of this gene. This is in contrast to what has been found for the hypX gene in R. leguminosarum, a hoxX homolog that is believed to be transcribed from the promoter for the large hyp operon (28). Since the end of the hypE gene is only 9 bp from the start of hoxX, it is possible that important sequences for hoxXA transcription are located within hypE.…”

Section: Discussioncontrasting

confidence: 83%

“…This indicates that the HoxX protein has a role in processing the hydrogenase enzyme, not in the regulation of transcription. A hypX mutant of R. leguminosarum also has this phenotype, and a role in processing the hydrogenase enzyme has been proposed for this gene product (28).…”

Section: Resultsmentioning

confidence: 99%

“…However, except for a potential membrane-spanning domain and a histidine kinase domain (24), HoxX has no homology with other known sensors of the NtrB family, which is considered the paradigm of sensor proteins (10). A homolog of HoxX, HypX, has recently been identified in the pea symbiont Rhizobium leguminosarum; it contains sequences similar to N 10 -formyl tetrahydrofolate-dependent enzymes, which are involved in the transfer of one-carbon units, and sequences conserved in the enoyl-coenzyme A hydratase/ isomerase family of enzymes (28). HypX has been shown to have a role in posttranslational processing of the hydrogenase enzyme; a possible role in metallocluster assembly has been proposed (28).…”

mentioning

confidence: 99%

“…A homolog of HoxX, HypX, has recently been identified in the pea symbiont Rhizobium leguminosarum; it contains sequences similar to N 10 -formyl tetrahydrofolate-dependent enzymes, which are involved in the transfer of one-carbon units, and sequences conserved in the enoyl-coenzyme A hydratase/ isomerase family of enzymes (28). HypX has been shown to have a role in posttranslational processing of the hydrogenase enzyme; a possible role in metallocluster assembly has been proposed (28). The high degree of sequence similarity between the predicted HoxX and the predicted HypX suggests that HoxX has a similar (i.e., processing) role in B. japonicum.…”

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confidence: 99%

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Roles of HoxX and HoxA in biosynthesis of hydrogenase in Bradyrhizobium japonicum

Durmowicz

Maier

1997

J Bacteriol

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In-frame deletion mutagenesis was used to study the roles of two Bradyrhizobium japonicum proteins, HoxX and HoxA, in hydrogenase biosynthesis; based on their sequences, these proteins were previously proposed to be sensor and regulator proteins, respectively, of a two-component regulatory system necessary for hydrogenase transcription. Deletion of the hoxX gene resulted in a strain that expressed only 30 to 40% of wild-type hydrogenase activity. The inactive unprocessed form of the hydrogenase large subunit accumulated in this strain, indicating a role for HoxX in posttranslational processing of the hydrogenase enzyme but not in transcriptional regulation. Strains containing a deletion of the hoxA gene or a double mutation (hoxX and hoxA) did not exhibit any hydrogenase activity under free-living conditions, and extracts from these strains were inactive in gel retardation assays with a 158-bp fragment of the DNA region upstream of the hupSL operon. However, bacteroids from root nodules formed by all three mutant types (hoxX, hoxA, and hoxX hoxA) exhibited hydrogenase activity comparable to that of wild-type bacteroids. Bacteroid extracts from all of these strains, including the wild type, failed to cause a shift of the hydrogenase upstream region used in our assay. It was shown that HoxA is a DNA-binding transcriptional activator of hydrogenase structural gene expression under free-living conditions but not under symbiotic conditions. Although symbiotic hydrogenase expression is still 54 dependent, a transcriptional activator other than HoxA functions presumably upstream of the HoxA binding site.Bradyrhizobium japonicum, the slow-growing, nitrogen-fixing symbiont of the soybean plant, expresses a hydrogen uptake hydrogenase that oxidizes hydrogen into its constituent protons and electrons. During symbiosis, the nickel-containing hydrogenase is thought to help improve the efficiency of nitrogen fixation in root nodules by passing H 2 -derived electrons to the electron transport chain, thereby recycling energy lost during nitrogen fixation (21). In B. japonicum, hydrogenase activity can also be induced under free-living, microaerobic conditions. Most studies of hydrogenase expression in this organism have been done under these conditions (21).Studies of the regulation of hydrogenase expression have shown that the hydrogenase structural genes, hupS and hupL, are regulated at the transcriptional level by hydrogen, oxygen, and nickel (14). Subsequently, a 50-bp region from base Ϫ99 to base Ϫ149 upstream of the transcriptional start site that is necessary for this regulation was identified (16). In addition, the hydrogenase promoter is 54 dependent and also uses integration host factor for full induction (4). Because the same cis-acting genetic region is responsible for regulation by the three environmental components, it is thought that the three signals are somehow integrated and passed on to a single activating factor that then binds within the 50-bp regulatory region to affect transcription (15).Previous sequenci...

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Section: Discussioncontrasting

confidence: 83%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Roles of HoxX and HoxA in biosynthesis of hydrogenase in Bradyrhizobium japonicum

Durmowicz

Maier

1997

J Bacteriol

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“…A similar situation exists in the symbiotic nitrogenfixing bacterium Rhizobium leguminosarum biovar viciae, where hupSL expression takes place in bacteroids only (Palacios et al, 1990) and requires the presence of NifA, which probably binds at an UAS in the hupSL promoter (Brito et al, 1995a,b). Although hypX, a gene homologous to hoxX gene, is present downstream of hypE (Rey et al, 1996), there is no evidence for the presence of a functional hoxA homologue in R. leguminosarum bv viciae (Brito et al, 1995b).…”

Section: Discussionmentioning

confidence: 99%

HoxA is a transcriptional regulator for expression of the hup structural genes in free‐living Bradyrhizobium japonicum

Soom

Wilde

Vanderleyden

1997

Molecular Microbiology

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SummaryA chromosomally integrated Bradyrhizobium japonicum hoxA mutant is unable to oxidize hydrogen in free-living conditions. Derepressing conditions that induce hydrogenase activity in free-living, wild-type B. japonicum cells cannot induce expression of the hydrogenase structural genes in the hoxA mutant. The DNA-binding capacity of HoxA at the hup promoter region was studied by means of gel retardation. Both heterotrophically growing cells and cells induced to express hydrogenase activity contain a protein that specifically binds to the hup promoter region. Crude protein extracts isolated from a B. japonicum hoxA mutant do not contain this binding compound. The HoxA protein was overexpressed in E. coli and isolated in the form of a maltose-binding protein (MBP)-HoxA fusion. The MBP-HoxA hybrid protein specifically bound to a 50 bp region of the hupSL promoter known to be important for regulation of hupSL expression.

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Nickel-binding proteins

Wattt

Ludden

1999

Cellular and Molecular Life Sciences (CMLS)

107

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Nickel enzymes are a relatively new class of metalloenzymes. The seven known nickel enzymes are urease, hydrogenase, CO-dehydrogenase, methyl-coenzyme M reductase, Ni-superoxide dismutase, glyoxalase I and cis-trans isomerase. The requirement for nickel implies the presence of a nickel-processing system, since free transition metals are harmful to the cell. A nickel-processing system involves the recognition and transport of nickel into the cell and the handling of the nickel once it enters the cell until it is inserted into the nickel enzyme. Several mechanisms for nickel transport have been identified and will be reviewed here. Accessory proteins required for the biosynthesis of the nickel active site have been identified. Accessory proteins bind the nickel when it enters the cell and are proposed to assist with the insertion of nickel into the enzyme. The function of the characterized nickel-processing proteins is described, and models for nickel insertion into the nickel enzymes are presented.

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The hydrogenase gene cluster ofRhizobium leguminosarum bv.viciae contains an additional gene (hypX), which encodes a protein with sequence similarity to the N10-formyltetrahydrofolate-dependent enzyme family and is required for nickel-dependent hydrogenase processing and activity

Cited by 32 publications

References 49 publications

Roles of HoxX and HoxA in biosynthesis of hydrogenase in Bradyrhizobium japonicum

Roles of HoxX and HoxA in biosynthesis of hydrogenase in Bradyrhizobium japonicum

HoxA is a transcriptional regulator for expression of the hup structural genes in free‐living Bradyrhizobium japonicum

Nickel-binding proteins

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