RESUMO
The 4 202 353 bp genome of the alkaliphilic bacterium Bacillus halodurans C-125 contains 4066 predicted protein coding sequences (CDSs), 2141 (52.7%) of which have functional assignments, 1182 (29%) of which are conserved CDSs with unknown function and 743 (18. 3%) of which have no match to any protein database. Among the total CDSs, 8.8% match sequences of proteins found only in Bacillus subtilis and 66.7% are widely conserved in comparison with the proteins of various organisms, including B.subtilis. The B. halodurans genome contains 112 transposase genes, indicating that transposases have played an important evolutionary role in horizontal gene transfer and also in internal genetic rearrangement in the genome. Strain C-125 lacks some of the necessary genes for competence, such as comS, srfA and rapC, supporting the fact that competence has not been demonstrated experimentally in C-125. There is no paralog of tupA, encoding teichuronopeptide, which contributes to alkaliphily, in the C-125 genome and an ortholog of tupA cannot be found in the B.subtilis genome. Out of 11 sigma factors which belong to the extracytoplasmic function family, 10 are unique to B. halodurans, suggesting that they may have a role in the special mechanism of adaptation to an alkaline environment.
Assuntos
Bacillus subtilis/genética , Bacillus/genética , Genoma Bacteriano , Transportadores de Cassetes de Ligação de ATP/genética , Adaptação Fisiológica/genética , Álcalis/metabolismo , Bacillus/química , Bacillus/classificação , Bacillus/metabolismo , Bacillus subtilis/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/fisiologia , Composição de Bases , Transporte Biológico , Parede Celular/genética , Parede Celular/metabolismo , Sequência Conservada/genética , Elementos de DNA Transponíveis/genética , Bases de Dados como Assunto , Metabolismo Energético , Evolução Molecular , Transferência Genética Horizontal/genética , Genes Bacterianos/genética , Genes Bacterianos/fisiologia , Dados de Sequência Molecular , Fases de Leitura Aberta/genética , Mapeamento Físico do Cromossomo , Biossíntese de Proteínas , RNA de Transferência/genética , Origem de Replicação/genética , Homologia de Sequência , Fator sigma/genética , Esporos Bacterianos/genética , Transcrição Gênica/genética , Transposases/genéticaRESUMO
The ribosomal RNA operons (rrn) of alkaliphilic Bacillus halodurans C-125 were characterized and compared with those of B. subtilis. We isolated clones containing rrn operons from a lambda phage library of the C-125 chromosome, and the complete nucleotide sequence of each was determined. Eight rrn operons were identified by PFGE analysis of the C-125 chromosome digested with I-CeuI. The transcriptional orientation of the rrn operons mapped on the chromosome by Southern hybridization analysis was the same as the direction of replication of the chromosome. These operons were designated as rrnA-H, starting from the oriC locus in clockwise rotation. Sequence and structural analyses of these operons suggested that six of the rrn operons in the C-125 chromosome, rrnA, rrnB, rrnC-rrnD, rrnE, and rrnH, correspond to rrnO, rrnA, rrnJ-rrnW, rrnI, and rrnD in B. subtilis, whereas the other rrn operons (rrnF and rrnG) were specifically observed in C-125. The rrn loci were positioned from 0 degrees to 90 degrees on the physical map, with the oriC locus assigned the position zero degrees. Two ORFs annotated as tnpA and ykfC, whose gene products are likely to act as transposases, were found downstream of these six operons. Comparative analysis of the 16S-23S and 23S-5S ITS (internally transcribed sequence) regions of B. halodurans C-125 and those of B. subtilis revealed that the ITS regions in C-125 were much longer than those in B. subtilis. There was no substantial difference in the length of potential promoter sequences in B. halodurans and B. subtilis.
Assuntos
Bacillus/genética , DNA Bacteriano/genética , Óperon de RNAr/genética , Álcalis , Bacillus subtilis/genética , Mapeamento Cromossômico , Clonagem Molecular , Eletroforese em Gel de Campo Pulsado , Endodesoxirribonucleases/metabolismo , Biblioteca Gênica , Concentração de Íons de Hidrogênio , Fases de Leitura Aberta/genética , Regiões Promotoras Genéticas/genética , RNA Bacteriano/análise , RNA Bacteriano/genética , RNA Mensageiro/análise , RNA Mensageiro/genéticaRESUMO
Seventeen Sse8387I linking clones isolated from the chromosome of Bacillus halodurans C-125 for the purpose of constructing a physical map were sequenced and analyzed by comparison with the BSORF database and the nonredundant protein databank. The orientations of Sse8387I or AscI linking clones serving to join adjacent fragments were determined by southern blot analysis using specific DNA probes. One-third of the open reading frames (ORFs) identified in the Sse8387I linking clones showed no significant similarity to any protein so far reported. The ORFs showing significant similarities to those of Bacillus subtilis were mapped in the chromosome of strain C-125, and the locations of the putative genes on the map were not well conserved between B. halodurans C-125 and B. subtilis.
Assuntos
Bacillus/genética , Cromossomos Bacterianos , Sequência de Bases , Clonagem Molecular , DNA Bacteriano , Mapeamento Físico do CromossomoRESUMO
Forty-one open reading frames (ORFs) were identified in a 32-kb DNA fragment of alkaliphilic Bacillus sp. C-125. A similarity search using the BSORF database found 37 ORFs with significant sequence similarity to B. subtilis RNA polymerase subunits, elongation factor G, elongation factor Tu, and ribosomal proteins. Each ORF product showed more than 70% identity to those of B. subtilis. Gene organization in the region of str, S10, spc, and the alpha cluster was highly conserved among three strains, C-125, B. subtilis, and B. stearothermophilus.
Assuntos
Bacillus/genética , Mapeamento Cromossômico , Família Multigênica , Fases de Leitura Aberta/genética , Proteínas Ribossômicas/genética , Bacillus/química , Sequência de Bases , Clonagem Molecular , Sequência Conservada/genética , Primers do DNA/química , DNA Bacteriano/química , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Análise de Sequência de DNARESUMO
Among alkaliphilic bacteria reported so far, Bacillus sp. C-125 is the strain most thoroughly characterized physiologically, biochemically, and genetically. A physical map of the chromosome of this strain was constructed to facilitate further genome analysis, and the genome size was revised from 3.7 to 4.25Mb. Complete digestion of the chromosomal DNA with two rare cut restriction endonucleases, AscI and Sse8387I, each yielded 20 fragments ranging in size from 20 to 600 kb. Seventeen linking clones were isolated in each instance to join the adjacent AscI or Sse8387I fragments in the chromosomal map. All AscI linking clones isolated were sequenced and analyzed by comparison with the BSORF database to map the genes in the chromosome of strain C-125. Several ORFs showing significant similarities to those of B. subtilis in the AscI linking clones were positioned on the physical map. The oriC region of the C-125 chromosome was identified by southern blot analysis with a DNA probe containing the gyrB region.
Assuntos
Bacillus/genética , Genoma Bacteriano , Mapeamento Físico do Cromossomo , Sequência de Bases , Clonagem Molecular , Primers do DNARESUMO
The nucleotide sequences of three independent fragments (designated no. 3, 4, and 9; each 15-20 kb in size) of the genome of alkaliphilic Bacillus sp. C-125 cloned in a lambda phage vector have been determined. Thirteen putative open reading frames (ORFs) were identified in sequenced fragment no. 3 and 11 ORFs were identified in no. 4. Twenty ORFs were also identified in fragment no. 9. All putative ORFs were analyzed in comparison with the BSORF database and non-redundant protein databases. The functions of 5 ORFs in fragment no. 3 and 3 ORFs in fragment no. 4 were suggested by their significant similarities to known proteins in the database. Among the 20 ORFs in fragment no. 9, the functions of 11 ORFs were similarly suggested. Most of the annotated ORFs in the DNA fragments of the genome of alkaliphilic Bacillus sp. C-125 were conserved in the Bacillus subtilis genome. The organization of ORFs in the genome of strain C-125 was found to differ from the order of genes in the chromosome of B. subtilis, although some gene clusters (ydh, yqi, yer, and yts) were conserved as operon units the same as in B. subtilis.
