RESUMO
Our approach to the isolation of plant-inducible bacterial genes of Azospirillum brasilense, based on the analysis of protein patterns of bacteria grown in the presence and in the absence of plant root exudates, led to the identification of an acidic 40 kDa protein. Cloning and sequencing analysis of the corresponding coding DNA region revealed the presence of two open reading frames transcribed in the same orientation. The deduced ORF1 protein, which corresponds to the 40 kDa protein, is very similar to the periplasmic ChvE protein, identified in Agrobacterium tumefaciens and involved in enhanced virulence. The deduced ORF2 protein shows homology to members of the LysR family of transcriptional regulators. The function of the ChvE-like protein in A. brasilense was investigated further. The protein, designated as SbpA (sugar binding protein A), is involved in the uptake of D-galactose and functions in the chemotaxis of A. brasilense towards several sugars, including D-galactose, L-arabinose and D-fucose. Expression of the sbpA gene requires the presence of the same sugars in the growth medium and is enhanced further in combination with carbon starvation of A. brasilense cells.
Assuntos
Azospirillum brasilense/genética , Metabolismo dos Carboidratos , Proteínas de Membrana Transportadoras , Proteínas de Transporte de Monossacarídeos/genética , Proteínas Periplásmicas de Ligação , Agrobacterium tumefaciens/química , Sequência de Aminoácidos , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Quimiotaxia/genética , Clonagem Molecular , Galactose/metabolismo , Regulação Bacteriana da Expressão Gênica , Dados de Sequência Molecular , Proteínas de Transporte de Monossacarídeos/química , Proteínas de Transporte de Monossacarídeos/metabolismo , Fases de Leitura Aberta , Raízes de Plantas/microbiologia , Ligação Proteica , Alinhamento de Sequência , Triticum/metabolismoRESUMO
Sequence analysis of a fragment of Azospirillum brasilense DNA revealed the presence of a ribA homologue, of which the 3' portion encodes a putative GTP cyclohydrolase II. The 5' portion (approx. half of the ORF) does not show homology to any other sequence from the databases.
Assuntos
Azospirillum brasilense/genética , Proteínas de Bactérias/genética , GTP Cicloidrolase/genética , Genes Bacterianos , Sequência de Aminoácidos , Azospirillum brasilense/enzimologia , Bacillus subtilis/química , Bacillus subtilis/enzimologia , Clonagem Molecular , Escherichia coli/química , Escherichia coli/enzimologia , Dados de Sequência Molecular , Alinhamento de Sequência , Homologia de Sequência de AminoácidosRESUMO
We have constructed a cosmid library of the Azospirillum brasilense Sp7 90-MDa plasmid (p90) and established the EcoRI restriction map of this plasmid. The central regions of cloned p90 DNA fragments from several recombinant cosmids were deleted by restriction endonuclease digestion and replaced by a DNA cassette encoding kanamycin resistance. Using these in vitro constructed deletions for marker exchange in Sp7, we made six different p90 deletion derivatives spanning all together 50% of the total length of p90. Comparison of the deletion derivatives with Sp7 for several properties revealed p90 loci involved in colony morphology, growth on minimal medium, motility, and adsorption to wheat roots. In analogy with the rhizobial symbiotic plasmids (pSym), we propose to denote the p90 plasmid as a rhizocoenotic plasmid (pRhico), carrying several genes involved in the A. brasilense-plant root interaction.