Analysis of the hypervariable region of the Salmonella enterica genome associated with tRNA(leuX)

The divergence of Salmonella enterica and Escherichia coli is estimated to have occurred approximately 140 million years ago. Despite this evolutionary distance, the genomes of these two species still share extensive synteny and homology. However, there are significant differences between the two species in terms of genes putatively acquired via various horizontal transfer events. Here we report on the composition and distribution across the Salmonella genus of a chromosomal region designated SPI-10 in Salmonella enterica serovar Typhi and located adjacent to tRNA(leuX). We find that across the Salmonella genus the tRNA(leuX) region is a hypervariable hot spot for horizontal gene transfer; different isolates from the same S. enterica serovar can exhibit significant variation in this region. Many P4 phage, plasmid, and transposable element-associated genes are found adjacent to tRNA(leuX) in both Salmonella and E. coli, suggesting that these mobile genetic elements have played a major role in driving the variability of this region.     

Homology between the invertible deoxyribonucleic acid sequence that controls flagellar-phase variation in Salmonella sp. and deoxyribonucleic acid sequences in other organisms.

The invertible deoxyribonucleic acid (DNA) segment cloned from Salmonella sp. was radioactively labeled and used as a probe to search for homologous sequences by Southern hybridization. Only one copy of the invertible segment could be found on the Salmonella sp. genome. Partial sequence homology with the invertible region was detected in bacteriophage Mu and P1 DNA by low-stringency hybridization. Under these conditions, no homology was detected with Escherichia coli DNA. A strain of Salmonella sp. defective in phase variation carrying the vH2- allele was also analyzed by DNA-DNA hybridization. The results show that there is sequence divergence between diphasic and vH2- strains within the invertible sequence.     

Structural homologies among type I restriction-modification systems.

Structural homologies among different restriction systems of Escherichia coli and several Salmonella species have been investigated by immunological methods using antibodies prepared against two subunits of the E. coli K12 restriction enzyme, and by DNA hybridization experiments using different fragments of the E. coli K12 hsd genes as probes. The results with both techniques show a strong homology between the E. coli K12 and B restriction-modification systems, weaker but nevertheless marked homology between E. coli K12 and the Salmonella systems SB, SP, and SQ and, surprisingly, no homology between the E. coli K12 and A systems.     

Sequence of the phosphomannose isomerase-encoding gene of Salmonella typhimurium

The pmi gene, encoding phosphomannose isomerase, of Salmonella typhimurium, was cloned in Escherichia coli K-12, and the protein product visualised in minicells. The cloned gene was sequenced; there was 77.4% nucleotide homology between the cloned pmi gene and the analogous manA gene of E. coli K-12, and 86.2% amino acid sequence homology between their presumptive gene products.     

DNA sequencing of a pathogenicity-related plasmid of an avian septicemic Escherichia coli strain

A 43-MDa conjugative plasmid isolated from an avian septicemic Escherichia coli (APEC) strain possessing genes related to the adhesion and invasion capacities of in vitro-cultured cells was sequenced. The results demonstrated that the 43-MDa plasmid harbors bacterial pathogenicity-related sequences which probably allow the wild-type pathogenic strain to adhere to and invade tissues and to cause septicemia in poultry. The existence of homology sequences to sequences belonging to other human pathogenic Enterobacteriaceae like Escherichia coli O157:H7, Shigella and Salmonella was also observed. The presence of these sequences in this plasmid could indicate that there is horizontal genetic transfer between bacterial strains isolated from different host species. In conclusion, the present study suggests that APEC strains harbor high-molecular weight plasmids that present pathogenicity-related sequences and that these are probably responsible for the pathogenicity exhibited by these strains. The presence of human pathogenicity-associated sequences in APEC conjugative plasmids suggests that these strains could represent a zoonotic risk.     

Molecular cloning and physical and functional characterization of the Salmonella typhimurium and Salmonella typhi galactose utilization operons.

The chromosomally encoded galactose utilization (gal) operons of Salmonella typhimurium and S. typhi were each cloned on similar 5.5-kilobase HindIII fragments into pBR322 and were identified by complementation of Gal- Escherichia coli strains. Restriction endonuclease analyses indicated that these Salmonellae operons share considerable homology, but some heterogeneities in restriction sites were observed. Subcloning and exonuclease mapping experiments showed that both operons have the same genetic organization as that established for the E. coli gal operon (i.e., 5' end, promoter, epimerase, transferase, kinase, and 3' end). Two gal operator regions (oE and oI) of S. typhimurium, identified by repressor titration in an E. coli superrepressor [galR(Sup)] mutant, were sequenced and found to flank the promoter region. This promoter region is identical to the -10 and -35 regions of the E. coli gal operon. Minicell studies demonstrated that the three gal structural genes of S. typhimurium encode separate polypeptides of 39 kilodaltons (kDa) (epimerase, 337 amino acids [aa's]), 41 kDa (transferase, 348 aa's), and 43 kDa (kinase, 380 aa's). Despite functional and organizational similarities, DNA sequence analysis revealed that the S. typhimurium gal genes show less than 70% homology to the E. coli gal operon. Because of codon degeneracy, the deduced amino acid sequences of these polypeptides are highly conserved (greater than 90% homology) as compared with those of the E. coli gal enzymes. These studies have defined basic genetic parameters of the gal genes of two medically important Salmonella species, and our findings support the hypothesized divergent evolution of E. coli and Salmonella spp. from a common ancestral parent bacterium.     

Map of DNA homology between the genomes of Salmonella bacteriophages P22 and L.

The genomes of temperate Salmonella typhimurium phages P22 and L share approximately 69% homology, as measured by DNA heteroduplex analysis. Alignment of the P22/L heteroduplex molecules with a P22 physical map places most of this homology between the capsid genes and genes in the vicinity of the prophage attachment sites. The degree of genetic relatedness between these phages and the lambdoid phages is also discussed.     

Three-dimensional structural model of the serine receptor ligand-binding domain.

Computer-based homology modeling techniques were used to construct a three-dimensional model of the Escherichia coli serine receptor ligand-binding domain based on the crystal structure of the Salmonella typhimurium aspartate receptor and the sequence homology between the two receptors. Residues that have been found in mutagenesis studies to be necessary for serine binding are located in a proposed serine-binding site. Several other mutations that affect swimming behavior require relatively small shifts in alpha-carbon positions in the model to give a minimized structure, suggesting that small changes in receptor conformation can affect the signaling state of the receptor.     

The araBAD operon of Salmonella typhimurium LT2. I. Nucleotide sequence of araB and primary structure of its product, ribulokinase

Hybrid plasmids containing the araBAD operon of Salmonella typhimurium LT2 were characterized by Southern blot and genetic analyses. The nucleotide sequence of araB was determined. The araB gene product, ribulokinase (EC 2.7.1.16), was purified and the results of amino acid composition analysis and partial amino acid sequence are in agreement with predictions from the DNA sequence. Ribulokinase is 569 amino acid residues long and has a calculated Mr of 61 793. Ribulokinase shares significant homology with xylulose kinase from Escherichia coli. Codon usage in the araB gene does not favor those codons which have intermediate codon-anticodon binding energy.     

Molecular cloning and characterization of supQ/newD, a gene substitution system for the leuD gene of Salmonella typhimurium.

The isopropylmalate isomerase of Salmonella typhimurium and Escherichia coli is a complex of the leuC and leuD gene products. The supQ/new D gene substitution system in S. typhimurium restores leucine prototrophy to leuD mutants of S. typhimurium. Previous genetic evidence supports a model that indicates the replacement of the missing LeuD polypeptide by the newD gene product. This model proposed that this gene substitution is possible when a mutation at the supQ locus (near newD) liberates unaltered newD polypeptide from its normal complex with the supQ protein product. In this study, recombinant plasmids carrying newD, supQ, or both were transformed into E. coli and S. typhimurium strains deleted for the leuD and supQ genes to test the supQ/newD gene substitution model for suppression of leucine auxotrophy. It was determined that the newD gene encodes a 22-kilodalton polypeptide which can restore leucine prototrophy to leuD deletion strains and that a functional supQ gene prevents this suppression. It was also determined that the supQ and newD genes are separated by a gene encoding a 50-kilodalton protein, pB. While there is extensive DNA sequence homology between the leucine operons of S. typhimurium and E. coli, DNA hybridization experiments did not indicate substantial homology between the newD and leuD genes. These data, taken together with previously obtained genetic data, eliminate the possibility that supQ and newD are recently translocated segments of the leucine operon.     

The ends of Tn10 are not IS3.

By heteroduplex and hybridization analysis we showed that the inverted repetition (here called IS10) at the ends of the translocatable tetracycline resistance element Tn10 is not IS3, as had previously been reported by Ptashne and Cohen (J. Bacteriol. 122:776--781, 1975). Further analysis confirmed the homology between IS3 and the alpha beta sequence of F and demonstrated that IS10 was not present in the genomes of Salmonella typhimurium LT2 or Escherichia coli K-12.     

Adenylate kinase isoenzymes in Aspergillus nidulans

Multiple HindIII-restriction fragments of Salmonella typhimurium and Salmonella typhi chromosomal DNA exhibited homology with the heat-labile enterotoxin (LT1) gene of Escherichia coli as determined by Southern blot analysis. A 9.4 kb HindIII restriction fragment identified in S. typhimurium and S. typhi chromosomal DNA reacted with both eltA and eltB gene probes. However, the homology of the 9.4 kb DNA fragment from these Salmonella species was greater with eltB than eltA. In addition, a synthetic oligonucleotide probe, made to a portion of the putative GM1-ganglioside binding region of cholera toxin (CT) and LT1, hybridized with the 9.4 kb DNA fragment of S. typhimurium but not with the 9.4 kb fragment found in S. typhi isolates. The hybridization of multiple restriction fragments of Salmonella DNA with eltA and eltB gene sequences further suggests duplication of the stx operon on the chromosome of these bacteria.     

Evidence for a DNA inversion system in Bordetella pertussis

The expression of virulence-associated genes in Bordetella pertussis can be lost in three ways: phase variation, antigenic modulation, or serotype conversion. The mechanism(s) of these alterations in gene expression is unclear. B. pertussis chromosomal DNA was probed with cloned pin genes from Escherichia coli and cloned hin genes from Salmonella typhimurium. DNA duplex melting temperature experiments indicated significant homology between B. Pertussis chromosomal DNA and both DNA inversion genes. Southern blots using the hin gene probe showed homology with a 15 kb EcoRI fragment of B. pertussis chromosomal DNA. We postulate here that B. pertussis contains a DNA inversion system which may be responsible for serotype conversion or virulence phase change in this organism.     

Cloning of a sequence of Aquaspirillum magnetotacticum that complements the aroD gene of Escherichia coli

A 2 kb DNA fragment isolated from a cosmid library of Aquaspirillum magnetotacticum strain MS-1 complements the aromatic-metabolite requirements and iron-uptake deficiencies of Escherichia coli and Salmonella typhimurium strains that lack a functional aroD (biosynthetic dehydrodquinase) sequence. All recombinant cosmids selected for their aroD complementation property carry this sequence. No DNA sequence homology has, however, been detected by Southern hybridization between the cloned fragment and the aroD gene of E. coli or the qa2 (catabolic dehydroquinase) gene of Neurospora crassa.     

Expression of 9 Salmonella typhimurium enzymes for cobinamide synthesis. Identification of the 11-methyl and 20-methyl transferases of corrin biosynthesis.

Nine of the cbi genes from the 17.5 kb cob operon of Salmonella typhimurium previously shown by genetic studies to be involved in the biosynthesis of cobinamide from precorrin-2, have been subcloned and expressed in Escherichia coli. Seven of the gene products were found in the soluble fraction of cell lysates and have been purified. The gene products corresponding to cbi E, F, H and L were shown by SAM binding and by homology with other SAM-binding proteins to be candidates for the methyltransferases of vitamin B12 biosynthesis. The enzymatic functions of the gene products of cbiL and cbiF are associated with C-methylation at C-20 of precorrin-2 and C-11 of precorrin-3.     

Characterization of a plasmid-encoded urease gene cluster found in members of the family Enterobacteriaceae.

Plasmid-encoded urease gene clusters found in uropathogenic isolates of Escherichia coli, Providencia stuartii, and Salmonella cubana demonstrated DNA homology, similar positions of restriction endonuclease cleavage sites, and manners of urease expression and therefore represent the same locus. DNA sequence analysis indicated that the plasmid-encoded urease genes are closely related to the Proteus mirabilis urease genes.     

Biosynthesis of bacterial glycogen: purification and structural properties of Rhodospirillum tenue adenosine diphosphate glucose synthetase.

Adenosine diphosphate glucose synthetase from the photosynthetic bacterium Rhodospirillum tenue has been purified greater than 95%. The molecular weight of the enzyme is approximately 215,000, with a subunit molecular weight of about 51,000. The enzyme appears to be composed of four similar if not identical subunits. Although the amino acid composition of the enzyme is similar to that of Escherichia coli and Salmonella typhimurium, no apparent homology has been observed between their N-terminal amino acid sequences. Antisera prepared against the R. tenue enzyme can partially inhibit the activities of adenosine diphosphate glucose synthetases from other photosynthetic bacteria.