Primary Structure of the DNA-Binding Protein HRm from Rhizobium meliloti

Lainé, Bernard; Bélaïche, Denise; Khanaka, Hussain; Sáutière, Pierre

doi:10.1111/j.1432-1033.1983.tb07265.x

Cited by 42 publications

(14 citation statements)

References 27 publications

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“…AF042348). Located 221 bp downstream from the lon gene, a second ORF was identified in the same orientation having 100% inferred amino acid identity to HupB (results not shown), a histone-like protein HU subunit previously sequenced from S. meliloti (29). This close association of lon and hupB has also been noted in other organisms (7,50).…”

Section: Resultsmentioning

confidence: 61%

The Sinorhizobium meliloti Lon Protease Is Involved in Regulating Exopolysaccharide Synthesis and Is Required for Nodulation of Alfalfa

Summers¹,

Botero²,

Busse

et al. 2000

J Bacteriol

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While screening for Sinorhizobium meliloti Pho regulatory mutants, a transposon mutant was isolated that constitutively expressed higher levels of acid and alkaline phosphatase enzymes. This mutant was also found to form pseudonodules on alfalfa that were delayed in appearance relative to those formed by the wild-type strain, it contained few bacteroids, and it did not fix nitrogen. Sequence analysis of the transposon insertion site revealed the affected gene to have high homology to Lon proteases from a number of organisms. In minimal succinate medium, the mutant strain was found to grow more slowly, reach lower maximal optical density, and produce more extracellular polysaccharide (EPS) than the wild-type strain. The mutant fluoresced brightly on minimal succinate agar containing calcofluor (which binds to EPSI, a constitutively expressed succinoglycan), and gas chromotographic analysis of purified total EPS showed that the glucose-to-galactose ratio in the lon mutant total EPS was 5.0 ؎ 0.2 (mean ؎ standard error), whereas the glucose-to-galactose ratio in the wild-type strain was 7.1 ؎ 0.5. These data suggested that in addition to EPSI, the lon mutant also constitutively synthesized EPSII, a galactoglucan which is the second major EPS known to be produced by S. meliloti, but typically is expressed only under conditions of phosphate limitation. 13 C nuclear magnetic resonance analysis showed no major differences between EPS purified from the mutant and wild-type strains. Normal growth, EPS production, and the symbiotic phenotype were restored in the mutant strain when the wild-type lon gene was present in trans. The results of this study suggest that the S. meliloti Lon protease is important for controlling turnover of a constitutively expressed protein(s) that, when unregulated, disrupts normal nodule formation and normal growth.

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Section: Resultsmentioning

confidence: 61%

The Sinorhizobium meliloti Lon Protease Is Involved in Regulating Exopolysaccharide Synthesis and Is Required for Nodulation of Alfalfa

Summers¹,

Botero²,

Busse

et al. 2000

J Bacteriol

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“…The amino acid Strategy used for the determination of the amino acid sequence ofproteins HAt, HRl18 and HRI,, The tryptic peptides generated from each protein were characterized by their behaviour on reverse-phase high-pressure liquid chromatography, their amino acid composition and their amino acid sequences. As they were identical or showed very slight differences with those of protein HRm, they were aligned by comparison with the amino acid sequence of protein HRm which has been completely determined [18]. Furthermore the amino-terminal sequence of protein HAt was confirmed by the data provided by automated sequence analysis of the native molecule.…”

Section: Resultsmentioning

confidence: 99%

“…cleaved with a low yield and the peptide T-7 covering the sequence 60 -74 was obtained predominantly with respect to the peptide which covers the sequence 62-74. In protein HRm [18] this bond was also found resistant to both trypsin and protease from the submaxillary gland of mice. On the other hand, digestion of protein HAt with carboxypeptidase A for 2 h released the following residues (mol/mol protein) : serine 0.9, asparagine 0.9, valine 0.8 and alanine 0.7.…”

Section: Amino-acid Sequence Of Protein Hatmentioning

confidence: 97%

“…Alternatively peptides were separated by chromatography or electrophoresis on thin-layer sheets (Polygram cel300 from Macherey-Nagel, FRG) using solvent or buffer systems described in [26]. Amino-acid analyses of proteins and peptides were performed as in [18].…”

Section: Fractionation Of Pep T Idesmentioning

confidence: 99%

“…Acetonitrile for reverse-phase high-pressure liquid chromatography was from Carlo Erba. All solvents and reagents for sequence determination were as indicated in [18]. …”

mentioning

confidence: 99%

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Characterization and primary structures of DNA‐binding HU‐type proteins from Rhizobiaceae

Khanaka

Lainé

Sáutière

et al. 1985

European Journal of Biochemistry

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The DNA-binding HU-type proteins from several species of Rhizobiaceae including Rhizobium meliloti, two strains of Rhizobium leguminosarum with highly different phenotypic characters and Agrobacterium tumefaciens, were characterized and their amino acid sequences were determined.HU-type proteins isolated from R. leguminosarum L18 and A . tumefaciens are identical and show slight differences with the R. [5] and can fold up circular double-stranded DNA into nucleosomelike structures [6]. This protein, which is associated with the chromosomal and extrachromosomal DNA [7,8], is assumed to be involved in the condensation of the DNA. Furthermore protein HU stimulates the enzymatic replication of the chromosomal origin of E. coli [9] and modulates transcription [l, 101. HU-type proteins have been isolated from a variety of bacterial families [ l l -171 and sequence analyses of these proteins show that they are strongly conserved [18, 191 when compared to other prokaryotic proteins such as cytochromes [20]. Our aim is to investigate the extent of amino acid sequence conservation of HU-type proteins within the family Rhizobiaceae which has been shown to be highly heterogenous [21]. The current classification based upon host infectivity recognizes six species which are distributed into two biochemically very different groups : the slowgrowing Rhizobiaceae and the fast-growing Rhizobiaceae in which the genus Agrobacterium is included because of their close taxonomic relationship. Previous work showed that the physico-chemical properties of the HU-type protein isolated from the slow-growing R. japonicum differ markedly from those of homologous proteins from fast-growing Rhizobiaceae [22]. This paper deals with the comparison of the physicochemical and structural properties of HU-type proteins from fast-growing Rhizobiaceae. On the basis of phenotypic characters, several species, such as Rhizobium meliloti and Agrobacterium, appear homogeneous whereas others are hetAbbreviations. HPLC, high-pressure liquid chromatography; HU, heat-stable protein isolated from Escherichia coli strain U ; PhMeS02F, phenylmethylsulfonyl fluoride; TosPheCH2C1, N-tosylphenylalanyl chloromethane.Enzymes. Carboxypeptidase A (EC 3.4.17.1); trypsin (EC 3.4.21.4).erogeneous [23]. One strain of R. meliloti and Agrobacterium tumefaciens and two strains of R . leguminosarum displaying highly different phenotypic characters were chosen for our study. MATERIALS AND METHODS MaterialsRhizobium meliloti strain 2011 str. 3 (from Institut National de la Recherche Agronomique, Versailles, France), Agrobacterium tumefaciens strain B 6 (from Rijksuniversiteit, Gent, Belgium), Rhizobium leguminosarum strains L18 and LS3 (from Research Institute of Ontario, Canada) were grown as described in [24].Carboxypeptidase A (treated with PhMeS02F) was obtained from Worthington. Acetonitrile for reverse-phase high-pressure liquid chromatography was from Carlo Erba. All solvents and reagents for sequence determination were as indicated in [18]. Nomenclature of proteinsHU-type...

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Electron microscopic study of DNA complexes with proteins from the Archaebacterium Sulfolobus acidocaldarius

et al. 1986

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DNA‐protein complexes formed in vitro with isolated DNA‐binding proteins from the thermoacidophilic archaebacterium Sulfolobus acidocaldarius were analyzed by electron microscopy. Two of the proteins (10a and 10b) formed specific structures after incubation with DNA. Protein 10a, at low protein concentrations, showed individual small spots on the DNA and at high concentrations evenly covered doublestranded DNA. Protein 10b showed three different types of regular structures: one with single‐stranded and two with double‐stranded DNA. Using double‐stranded DNA, 10b first bound cooperatively to two strands forming long, plait‐like structures only slightly shorter than respective free DNA. The complex consists of two right‐handed, interwound fibers, with a helical pitch of 26 nm and a diameter of ∼10‐11 nm. At higher protein concentration than needed to package all DNA into the complex with two double‐stranded DNAs, the two DNAs were separated again and a new structure was formed evenly covering only one DNA strand. Both structures showed no significant contraction of the length of the DNA covered in the complex. Nucleoprotein formed with single‐stranded ΦX174 DNA had a diameter of ∼11 nm and could form circles with a contour length of 0.5 μm.

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Primary Structure of the DNA-Binding Protein HRm from Rhizobium meliloti

Cited by 42 publications

References 27 publications

The Sinorhizobium meliloti Lon Protease Is Involved in Regulating Exopolysaccharide Synthesis and Is Required for Nodulation of Alfalfa

The Sinorhizobium meliloti Lon Protease Is Involved in Regulating Exopolysaccharide Synthesis and Is Required for Nodulation of Alfalfa

Characterization and primary structures of DNA‐binding HU‐type proteins from Rhizobiaceae

Electron microscopic study of DNA complexes with proteins from the Archaebacterium Sulfolobus acidocaldarius

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