| 1. |
- Eisenhut, Marion, et al.
(författare)
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The plant-like C2 glycolate cycle and the bacterial-like glycerate pathway cooperate in phosphoglycolate metabolism in cyanobacteria.
- 2006
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Ingår i: Plant Physiology. - : Oxford University Press (OUP). - 0032-0889 .- 1532-2548. ; 142:1
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Tidskriftsartikel (refereegranskat)abstract
- The occurrence of a photorespiratory 2-phosphoglycolate metabolism in cyanobacteria is not clear. In the genome of the cyanobacterium Synechocystis sp. strain PCC 6803, we have identified open reading frames encoding enzymes homologous to those forming the plant-like C2 cycle and the bacterial-type glycerate pathway. To study the route and importance of 2-phosphoglycolate metabolism, the identified genes were systematically inactivated by mutagenesis. With a few exceptions, most of these genes could be inactivated without leading to a high-CO(2)-requiring phenotype. Biochemical characterization of recombinant proteins verified that Synechocystis harbors an active serine hydroxymethyltransferase, and, contrary to higher plants, expresses a glycolate dehydrogenase instead of an oxidase to convert glycolate to glyoxylate. The mutation of this enzymatic step, located prior to the branching of phosphoglycolate metabolism into the plant-like C2 cycle and the bacterial-like glycerate pathway, resulted in glycolate accumulation and a growth depression already at high CO(2). Similar growth inhibitions were found for a single mutant in the plant-type C2 cycle and more pronounced for a double mutant affected in both the C2 cycle and the glycerate pathway after cultivation at low CO(2). These results suggested that cyanobacteria metabolize phosphoglycolate by the cooperative action of the C2 cycle and the glycerate pathway. When exposed to low CO(2), glycine decarboxylase knockout mutants accumulated far more glycine and lysine than wild-type cells or mutants with inactivated glycerate pathway. This finding and the growth data imply a dominant, although not exclusive, role of the C2 route in cyanobacterial phosphoglycolate metabolism.
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| 2. |
- Hagemann, Martin, et al.
(författare)
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Detection of a phage genome carrying a zonula occludens like toxin gene (zot) in clinical isolates of Stenotrophomonas maltophilia.
- 2006
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Ingår i: Archives of Microbiology. - : Springer Science and Business Media LLC. - 0302-8933 .- 1432-072X. ; 185:6
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Tidskriftsartikel (refereegranskat)abstract
- During a study of the genetic diversity of Stenotrophomonas strains, we found an autonomous replicating DNA molecule in chromosomal DNA preparations of the clinical Stenotrophomonas maltophilia strain c5. The entire sequence of 6,907 bp of the isolated DNA molecule was determined, which was called phiSMA9. Seven ORFs, which code for proteins with considerable similarity to proteins in databases, were identified in the DNA sequence. The largest ORF shows high sequence similarities to the pI protein of the filamentous phage phiLf, which was later shown to be identical to toxin Zot of Vibrio cholerae. Beside the Zot-like protein, six other proteins with similarities to known phage proteins such as a phage replication protein RstA and phage absorption or coat protein are encoded on phiSMA9, which indicate that this circular DNA molecule represents the replicative form of a linear phage genome. A PCR-based screening showed that only five from the totally investigated 47 Stenotrophomonas strains of clinical and environmental origin harbor these genes. Altogether, we describe the first genome of a phage for the nosocomial pathogen Stenotrophomonas, which contains a Zot toxin like gene and might be regarded as the first Stenotrophomonas virulence factor.
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| 3. |
- Hagemann, Martin, et al.
(författare)
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The plant-associated bacterium Stenotrophomonas rhizophila expresses a new enzyme for the synthesis of the compatible solute glucosylglycerol.
- 2008
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Ingår i: Journal of Bacteriology. - 0021-9193 .- 1098-5530. ; 190:17
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Tidskriftsartikel (refereegranskat)abstract
- The rhizobacterium Stenotrophomonas rhizophila accumulates the compatible solutes glucosylglycerol (GG) and trehalose under salt stress conditions. The complete gene for the GG synthesis enzyme was cloned and sequenced. This enzyme from S. rhizophila represented a novel fusion protein composed of a putative C-terminal GG-phosphate synthase domain and an N-terminal putative GG-phosphate phosphatase domain, which was named GgpPS. A similar gene was cloned from Pseudomonas sp. strain OA146. The ggpPS gene was induced after a salt shock in S. rhizophila cells. After the salt-loaded cells reached stationary phase, the ggpPS mRNA content returned to the low level characteristic of the control cells, and GG was released into the medium. The complete ggpPS gene and a truncated version devoid of the phosphatase part were obtained as recombinant proteins. Enzyme activity tests revealed the expected abilities of the full-length protein to synthesize GG and the truncated GgpPS to synthesize GG-phosphate. However, dephosphorylation of GG-phosphate was detected only with the complete GgpPS protein. These enzyme activities were confirmed by complementation experiments using defined GG-defective mutants of the cyanobacterium Synechocystis sp. strain PCC 6803. Genes coding for proteins very similar to the newly identified fusion protein GgpPS for GG synthesis in S. rhizophila were found in genome sequences of related bacteria, where these genes are often linked to a gene coding for a transporter of the Mfs superfamily.
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| 4. |
- Hasse, Dirk, et al.
(författare)
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Alternative splicing produces an H-protein with better substrate properties for the P-protein of glycine decarboxylase.
- 2009
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Ingår i: The FEBS Journal. - : Wiley. - 1742-464X .- 1742-4658. ; 276:23
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Tidskriftsartikel (refereegranskat)abstract
- Several thousand plant genes are known to produce multiple transcripts, but the precise function of most of the alternatively encoded proteins is not known. Alternative splicing has been reported for the H-protein subunit of glycine decarboxylase in the genus Flaveria. H-protein has no catalytic activity itself but is a substrate of the three enzymatically active subunits, P-, T- and L-protein. In C(4) species of Flaveria, two H-proteins originate from single genes in an organ-dependent manner. Here, we report on differences between the two alternative H-protein variants with respect to their interaction with the glycine-decarboxylating subunit, P-protein. Steady-state kinetic analyses of the alternative Flaveria H-proteins and artificially produced 'alternative' Arabidopsis H-proteins, using either pea mitochondrial matrix extracts or recombinant cyanobacterial P-protein, consistently demonstrate that the alternative insertion of two alanine residues at the N-terminus of the H-protein elevates the activity of P-protein by 20%in vitro, and could promote glycine decarboxylase activity in vivo.
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| 5. |
- Hasse, Dirk, et al.
(författare)
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Properties of recombinant glycine decarboxylase P- and H-protein subunits from the cyanobacterium Synechocystis sp. strain PCC 6803.
- 2007
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Ingår i: FEBS Letters. - : Wiley. - 0014-5793 .- 1873-3468. ; 581:7
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Tidskriftsartikel (refereegranskat)abstract
- The multi-enzyme complex glycine decarboxylase is important for one-carbon metabolism, essential for the photorespiratory glycolate cycle of plants, and comprises four different polypeptides, P-, H-, T-, and L-protein. We report on the production and properties of recombinant P-protein from the cyanobacterium Synechocystis and also describe features of recombinant H-protein from the same organism. The P-protein shows enzymatic activity with lipoylated H-protein and very low activity with H-apoprotein or lipoate as artificial cofactors. Its affinity towards glycine is unaffected by the presence and nature of the methyleneamine acceptor molecule. The cyanobacterial H-protein apparently forms stable dimers.
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| 6. |
- Ribbeck-Busch, Kathrin, et al.
(författare)
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A molecular biological protocol to distinguish potentially human pathogenic Stenotrophomonas maltophilia from plant-associated Stenotrophomonas rhizophila.
- 2005
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Ingår i: Environmental Microbiology. - : Wiley. - 1462-2912 .- 1462-2920. ; 7:11
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Tidskriftsartikel (refereegranskat)abstract
- In recent years, the importance of the Gram-negative bacterium Stenotrophomonas as an opportunistic pathogen as well as in biotechnology has increased. The aim of the present study was to develop new methods for distinguishing between strains closely related to the potentially human pathogenic Stenotrophomonas maltophilia and those closely related to the plant-associated Stenotrophomonas rhizophila. To accomplish this, 58 strains were characterized by 16S rDNA sequencing and amplified ribosomal DNA restriction analysis (ARDRA), and the occurrence of specific functional genes. Based on 16S rDNA sequences, an ARDRA protocol was developed which allowed differentiation between strains of the S. maltophilia and the S. rhizophila group. As it was known that only salt-treated cells of S. rhizophila were able to synthesize the compatible solute glucosylglycerol (GG), the ggpS gene responsible for GG synthesis was used for differentiation between both species and it was confirmed that it only occurred in S. rhizophila strains. As a further genetic marker the smeD gene, which is part of the genes coding for the multidrug efflux pump SmeDEF from S. maltophilia, was used. Based on the results we propose a combination of fingerprinting techniques using the 16S rDNA and the functional genes ggpS and smeD to distinguish both Stenotrophomonas species.
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