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Devine, E.Holmqvist, M.Stensjo, K.Lindblad, P.
Diversity and transcription of proteases involved in the maturation of hydrogenases in Nostoc punctiforme ATCC 29133 and Nostoc sp. strain PCC 7120.
BMC Microbiol. 2009 Mar 11;9:53. doi: 10.1186/1471-2180-9-53. PMID:19284580 | Abstract | MeSH | Related Articles | Sections | Gene Index |
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BACKGROUND: The last step in the maturation process of the large subunit of [NiFe]-hydrogenases is a proteolytic cleavage of the C-terminal by a hydrogenase specific protease. Contrary to other accessory proteins these hydrogenase proteases are believed to be specific whereby one type of hydrogenases specific protease only cleaves one type of hydrogenase. In cyanobacteria this is achieved by the gene product of either hupW or hoxW, specific for the uptake or the bidirectional hydrogenase respectively. The filamentous cyanobacteria Nostoc punctiforme ATCC 29133 and Nostoc sp strain PCC 7120 may contain a single uptake hydrogenase or both an uptake and a bidirectional hydrogenase respectively. RESULTS: In order to examine these proteases in cyanobacteria, transcriptional analyses were performed of hupW in Nostoc punctiforme ATCC 29133 and hupW and hoxW in Nostoc sp. strain PCC 7120. These studies revealed numerous transcriptional start points together with putative binding sites for NtcA (hupW) and LexA (hoxW). In order to investigate the diversity and specificity among hydrogeanse specific proteases we constructed a phylogenetic tree which revealed several subgroups that showed a striking resemblance to the subgroups previously described for [NiFe]-hydrogenases. Additionally the proteases specificity was also addressed by amino acid sequence analysis and protein-protein docking experiments with 3D-models derived from bioinformatic studies. These studies revealed a so called "HOXBOX"; an amino acid sequence specific for protease of Hox-type which might be involved in docking with the large subunit of the hydrogenase. CONCLUSION: Our findings suggest that the hydrogenase specific proteases are under similar regulatory control as the hydrogenases they cleave. The result from the phylogenetic study also indicates that the hydrogenase and the protease have co-evolved since ancient time and suggests that at least one major horizontal gene transfer has occurred. This co-evolution could be the result of a close interaction between the protease and the large subunit of the [NiFe]-hydrogenases, a theory supported by protein-protein docking experiments performed with 3D-models. Finally we present data that may explain the specificity seen among hydrogenase specific proteases, the so called "HOXBOX"; an amino acid sequence specific for proteases of Hox-type. This opens the door for more detailed studies of the specificity found among hydrogenase specific proteases and the structural properties behind it.
MeSH terms
1. Amino Acid Sequence
2. Bacterial Proteins/genetics/*metabolism
3. DNA, Bacterial/genetics
4. Endopeptidases/genetics/*metabolism
5. Gene Expression Regulation, Bacterial
6. Hydrogenase/genetics/*metabolism
7. Models, Molecular
8. Molecular Sequence Data
9. Nostoc/enzymology/*genetics
10. Phylogeny
11. Protein Structure, Tertiary
12. Sequence Alignment
13. Sequence Analysis, DNA
14. Substrate Specificity
15. Transcription Initiation Site
16. Transcription, Genetic
2
Wunschiers, R.Batur, M.Lindblad, P.
Presence and expression of hydrogenase specific C-terminal endopeptidases in cyanobacteria.
BMC Microbiol. 2003 May 7;3:8. PMID:12735794 | Abstract | MeSH | Related Articles | Sections | Gene Index |
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BACKGROUND: Hydrogenases catalyze the simplest of all chemical reactions: the reduction of protons to molecular hydrogen or vice versa. Cyanobacteria can express an uptake, a bidirectional or both NiFe-hydrogenases. Maturation of those depends on accessory proteins encoded by hyp-genes. The last maturation step involves the cleavage of a ca. 30 amino acid long peptide from the large subunit by a C-terminal endopeptidase. Until know, nothing is known about the maturation of cyanobacterial NiFe-hydrogenases. The availability of three complete cyanobacterial genome sequences from strains with either only the uptake (Nostoc punctiforme ATCC 29133/PCC 73102), only the bidirectional (Synechocystis PCC 6803) or both NiFe-hydrogenases (Anabaena PCC 7120) prompted us to mine these genomes for hydrogenase maturation related genes. In this communication we focus on the presence and the expression of the NiFe-hydrogenases and the corresponding C-terminal endopeptidases, in the three strains mentioned above. RESULTS: We identified genes encoding putative cyanobacterial hydrogenase specific C-terminal endopeptidases in all analyzed cyanobacterial genomes. The genes are not part of any known hydrogenase related gene cluster. The derived amino acid sequences show only low similarity (28-41%) to the well-analyzed hydrogenase specific C-terminal endopeptidase HybD from Escherichia coli, the crystal structure of which is known. However, computational secondary and tertiary structure modeling revealed the presence of conserved structural patterns around the highly conserved active site. Gene expression analysis shows that the endopeptidase encoding genes are expressed under both nitrogen-fixing and non-nitrogen-fixing conditions. CONCLUSION: Anabaena PCC 7120 possesses two NiFe-hydrogenases and two hydrogenase specific C-terminal endopeptidases but only one set of hyp-genes. Thus, in contrast to the Hyp-proteins, the C-terminal endopeptidases are the only known hydrogenase maturation factors that are specific. Therefore, in accordance with previous nomenclature, we propose the gene names hoxW and hupW for the bidirectional and uptake hydrogenase processing endopeptidases, respectively. Due to their constitutive expression we expect that, at least in cyanobacteria, the endopeptidases take over multiple functions.
MeSH terms
1. Amino Acid Sequence
2. Cyanobacteria/*enzymology/genetics
3. Endopeptidases/*biosynthesis/genetics
4. Genes, Bacterial
5. Hydrogenase/*biosynthesis/genetics
6. Molecular Sequence Data
7. Protein Structure, Tertiary
8. Sequence Homology, Amino Acid
3
Lindberg, P.Devine, E.Stensjo, K.Lindblad, P.
HupW protease specifically required for processing of the catalytic subunit of the uptake hydrogenase in the cyanobacterium Nostoc sp. strain PCC 7120.
Appl Environ Microbiol. 2012 Jan;78(1):273-6. doi: 10.1128/AEM.05957-11. Epub 2011 Oct 21. PMID:22020512 | Abstract | MeSH | Related Articles | Sections | Gene Index |
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The maturation process of [NiFe] hydrogenases includes a proteolytic cleavage of the large subunit. We constructed a mutant of Nostoc strain PCC 7120 in which hupW, encoding a putative hydrogenase-specific protease, is inactivated. Our results indicate that the protein product of hupW selectively cleaves the uptake hydrogenase in this cyanobacterium.
MeSH terms
1. Bacterial Proteins/genetics/*metabolism
2. Catalytic Domain
3. Endopeptidases/genetics/*metabolism
4. Gene Knockout Techniques
5. Nostoc/*enzymology/genetics
6. Oxidoreductases/*metabolism
4
Agervald, A.Stensjo, K.Holmqvist, M.Lindblad, P.
Transcription of the extended hyp-operon in Nostoc sp. strain PCC 7120.
BMC Microbiol. 2008 Apr 28;8:69. doi: 10.1186/1471-2180-8-69. PMID:18442387 | Abstract | MeSH | Related Articles | Sections | Gene Index |
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BACKGROUND: The maturation of hydrogenases into active enzymes is a complex process and e.g. a correctly assembled active site requires the involvement of at least seven proteins, encoded by hypABCDEF and a hydrogenase specific protease, encoded either by hupW or hoxW. The N2-fixing cyanobacterium Nostoc sp. strain PCC 7120 may contain both an uptake and a bidirectional hydrogenase. The present study addresses the presence and expression of hyp-genes in Nostoc sp. strain PCC 7120. RESULTS: RT-PCRs demonstrated that the six hyp-genes together with one ORF may be transcribed as a single operon. Transcriptional start points (TSPs) were identified 280 bp upstream from hypF and 445 bp upstream of hypC, respectively, demonstrating the existence of several transcripts. In addition, five upstream ORFs located in between hupSL, encoding the small and large subunits of the uptake hydrogenase, and the hyp-operon, and two downstream ORFs from the hyp-genes were shown to be part of the same transcript unit. A third TSP was identified 45 bp upstream of asr0689, the first of five ORFs in this operon. The ORFs are annotated as encoding unknown proteins, with the exception of alr0692 which is identified as a NifU-like protein. Orthologues of the four ORFs asr0689-alr0692, with a highly conserved genomic arrangement positioned between hupSL, and the hyp genes are found in several other N2-fixing cyanobacteria, but are absent in non N2-fixing cyanobacteria with only the bidirectional hydrogenase. Short conserved sequences were found in six intergenic regions of the extended hyp-operon, appearing between 11 and 79 times in the genome. CONCLUSION: This study demonstrated that five ORFs upstream of the hyp-gene cluster are co-transcribed with the hyp-genes, and identified three TSPs in the extended hyp-gene cluster in Nostoc sp. strain PCC 7120. This may indicate a function related to the assembly of a functional uptake hydrogenase, hypothetically in the assembly of the small subunit of the enzyme.
MeSH terms
1. Bacterial Proteins/genetics
2. Base Pairing
3. Chromosome Mapping
4. Conserved Sequence
5. DNA, Bacterial/analysis/genetics
6. Hydrogenase/*genetics
7. Nostoc/*enzymology/*genetics
8. Open Reading Frames
9. *Operon
10. Promoter Regions, Genetic
11. Tandem Repeat Sequences
12. Transcription Initiation Site
13. *Transcription, Genetic
5
Agervald, A.Zhang, X.Stensjo, K.Devine, E.Lindblad, P.
CalA, a cyanobacterial AbrB protein, interacts with the upstream region of hypC and acts as a repressor of its transcription in the cyanobacterium Nostoc sp. strain PCC 7120.
Appl Environ Microbiol. 2010 Feb;76(3):880-90. doi: 10.1128/AEM.02521-09. Epub 2009 Dec 18. PMID:20023111 | Abstract | MeSH | Related Articles | Sections | Gene Index |
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The filamentous, heterocystous, nitrogen-fixing cyanobacterium Nostoc sp. strain PCC 7120 may contain, depending on growth conditions, up to two hydrogenases directly involved in hydrogen metabolism. HypC is one out of at least seven auxiliary gene products required for synthesis of a functional hydrogenase, specifically involved in the maturation of the large subunit. In this study we present a protein, CalA (Alr0946 in the genome), belonging to the transcription regulator family AbrB, which in protein-DNA assays was found to interact with the upstream region of hypC. Transcriptional investigations showed that calA is cotranscribed with the downstream gene alr0947, which encodes a putative protease from the abortive infection superfamily, Abi. CalA was shown to interact specifically not only with the upstream region of hypC but also with its own upstream region, acting as a repressor on hypC. The bidirectional hydrogenase activity was significantly downregulated when CalA was overexpressed, demonstrating a correlation with the transcription factor, either direct or indirect. In silico studies showed that homologues to both CalA and Alr0947 are highly conserved proteins within cyanobacteria with very similar physical organizations of the corresponding structural genes. Possible functions of the cotranscribed downstream protein Alr0947 are presented. In addition, we present a three-dimensional (3D) model of the DNA binding domain of CalA and putative DNA binding mechanisms are discussed.
MeSH terms
1. Amino Acid Sequence
2. Bacterial Proteins/chemistry/genetics/*metabolism
3. Conserved Sequence/genetics
4. DNA, Bacterial/genetics/metabolism
5. Endopeptidases/genetics/metabolism
6. Gene Expression Regulation, Bacterial
7. Genes, Bacterial
8. Hydrogen/metabolism
9. Hydrogenase/genetics/metabolism
10. Molecular Sequence Data
11. Nitrogen Fixation/genetics
12. Nostoc/*genetics/metabolism
13. Promoter Regions, Genetic
14. Protein Structure, Tertiary/genetics
15. Repressor Proteins/genetics/*metabolism
16. Sequence Alignment
17. Sequence Analysis, DNA
18. Sequence Analysis, Protein
19. Sequence Homology, Amino Acid
20. Transcription Factors/genetics/metabolism
6
Ehira, S.Ohmori, M.Sato, N.
Genome-wide expression analysis of the responses to nitrogen deprivation in the heterocyst-forming cyanobacterium Anabaena sp. strain PCC 7120.
DNA Res. 2003 Jun 30;10(3):97-113. PMID:12886952 | Abstract | MeSH | Related Articles | Sections | Gene Index |
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A heterocyst is a terminally differentiated cell of cyanobacteria which is specialized in dinitrogen fixation. Heterocyst differentiation in Anabaena sp. strain PCC 7120 is triggered by deprivation of combined nitrogen in the medium. Although various genes that are upregulated during heterocyst differentiation have been reported, most studies to date were limited to individual or a small number of genes. We prepared microarrays in collaboration with other members of the Anabaena Genome Project. Here we report on the genome-wide expression analysis of the responses to nitrogen deprivation in Anabaena. Many unidentified genes, as well as previously known genes, were found to be upregulated by nitrogen deprivation at various time points. Three main profiles of gene expression were found: genes expressed transiently at an early stage (1-3 hr) of nitrogen deprivation, genes expressed transiently at a later stage (8 hr), and genes expressed when heterocysts are formed (24 hr). We also noted that many of the upregulated genes were physically clustered to form 'expressed islands' on the chromosome. Namely, large, continuous genomic regions containing many genes were upregulated in a coordinated manner. This suggests a mechanism of global regulation of gene expression that involves chromosomal structure, which is reminiscent of eukaryotic chromatin remodelling. The possible implications of this global regulation are discussed.
MeSH terms
1. Anabaena/genetics/*growth & development/metabolism
2. Bacterial Proteins/genetics/*metabolism
3. Culture Media
4. *Gene Expression Regulation, Bacterial
5. *Genome, Bacterial
6. Nitrogen/*metabolism
7. Nitrogen Fixation
8. *Oligonucleotide Array Sequence Analysis
7
Holmqvist, M.Lindberg, P.Agervald, A.Stensjo, K.Lindblad, P.
Transcript analysis of the extended hyp-operon in the cyanobacteria Nostoc sp. strain PCC 7120 and Nostoc punctiforme ATCC 29133.
BMC Res Notes. 2011 Jun 14;4:186. doi: 10.1186/1756-0500-4-186. PMID:21672234 | Abstract | MeSH | Related Articles | Sections | Gene Index |
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BACKGROUND: Cyanobacteria harbor two [NiFe]-type hydrogenases consisting of a large and a small subunit, the Hup- and Hox-hydrogenase, respectively. Insertion of ligands and correct folding of nickel-iron hydrogenases require assistance of accessory maturation proteins (encoded by the hyp-genes). The intergenic region between the structural genes encoding the uptake hydrogenase (hupSL) and the accessory maturation proteins (hyp genes) in the cyanobacteria Nostoc PCC 7120 and N. punctiforme were analysed using molecular methods. FINDINGS: The five ORFs, located in between the uptake hydrogenase structural genes and the hyp-genes, can form a transcript with the hyp-genes. An identical genomic localization of these ORFs are found in other filamentous, N2-fixing cyanobacterial strains. In N. punctiforme and Nostoc PCC 7120 the ORFs upstream of the hyp-genes showed similar transcript level profiles as hupS (hydrogenase structural gene), nifD (nitrogenase structural gene), hypC and hypF (accessory hydrogenase maturation genes) after nitrogen depletion. In silico analyzes showed that these ORFs in N. punctiforme harbor the same conserved regions as their homologues in Nostoc PCC 7120 and that they, like their homologues in Nostoc PCC 7120, can be transcribed together with the hyp-genes forming a larger extended hyp-operon. DNA binding studies showed interactions of the transcriptional regulators CalA and CalB to the promoter regions of the extended hyp-operon in N. punctiforme and Nostoc PCC 7120. CONCLUSIONS: The five ORFs upstream of the hyp-genes in several filamentous N2-fixing cyanobacteria have an identical genomic localization, in between the genes encoding the uptake hydrogenase and the maturation protein genes. In N. punctiforme and Nostoc PCC 7120 they are transcribed as one operon and may form transcripts together with the hyp-genes. The expression pattern of the five ORFs within the extended hyp-operon in both Nostoc punctiforme and Nostoc PCC 7120 is similar to the expression patterns of hupS, nifD, hypF and hypC. CalA, a known transcription factor, interacts with the promoter region between hupSL and the five ORFs in the extended hyp-operon in both Nostoc strains.
MeSH terms

2.146822268 [0.202167178, 0.347543297, 2.118629612, 2.121005767, 2.146738192]

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