Purification, cloning, and preliminary characterization of a Spiroplasma citri ribosomal protein with DNA binding capacity.
The rpsB-tsf-x operon of Spiroplasma citri encodes ribosomal protein S2 and elongation factor Ts, two components of the translational apparatus, and an unidentified X protein. A potential DNA-binding site (a 20-base pair (bp) inverted repeat sequence) is located at the 3' end of rpsB. Southwestern analysis of S. citri proteins, with a 30-bp double-stranded oligonucleotide probe (IRS), containing the 20-bp inverted repeat sequence and the genomic flanking sequences, detected an IRS-binding protein of 46 kDa (P46). P46 protein, which displays preferential affinity for the IRS, was purified from S. citri by a combination of affinity and gel filtration chromatographies. The native form of P46 seems to be homomultimeric as estimated by SDS-polyacrylamide gel electrophoresis analysis and gel filtration. A 3.5-kilobase pair S. citri DNA fragment comprising the P46 gene and flanking sequences was cloned and sequenced. Sequence analysis of this DNA fragment indicated that the P46 gene is located within the S10-spc operon of S. citri at the position of the gene coding for ribosomal protein L29 in the known S10-spc operons. The similarity between the N-terminal domain of P46 and the L29 ribosomal protein family and the presence of a 46-kDa IRS-binding protein in S. citri ribosomes indicated that P46 is the L29 ribosomal protein of S. citri. We suggest that P46 is a bifunctional protein with an L29 N-terminal domain and a C-terminal domain involved in IRS binding.
Le Dantec
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Laboratoire de Biologie Cellulaire et Moléculaire, Institut National de la Recherche Agronomique and Université Victor Segalen Bordeaux 2, 33883 Villenave d'Ornon Cedex, France.
Castroviejo
M
M
Bové
J M
JM
Saillard
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C
D001425:Bacterial Outer Membrane Proteins / Q000235:genetics; D001483:Base Sequence; D002846:Chromatography, Affinity; D002874:Chromosome Mapping; D002876:Chromosomes, Bacterial; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000737:chemistry* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D005810:Multigene Family; D015345:Oligonucleotide Probes; D009876:Operon; D010445:Peptide Elongation Factors / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D012091:Repetitive Sequences, Nucleic Acid; D012269:Ribosomal Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013149:Spiroplasma / Q000235:genetics* / Q000378:metabolism*
D001425:Bacterial Outer Membrane Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D015345:Oligonucleotide Probes; D010445:Peptide Elongation Factors; D011994:Recombinant Proteins; D012269:Ribosomal Proteins; C023512:elongation factor Ts; C030004:ribosomal protein S2; C028588:spiralin