The 90kb plasmid resident in Salmonella typhimurium confers increased virulence in mice by promoting the spread of infection after invasion of the intestinal epithelium. The nucleotide sequence of a 13.9kb segment of this plasmid known to encode an outer membrane protein related in sequence to components of fimbrial biosynthesis in enteric bacteria was determined. This cloned segment between the repB and repC replicon regions programmed expression of abundant surface fimbriae in Escherichia coli and S. typhimurium cells. A 7kb region contained seven open reading frames, the protein products of five of which were related in sequence to regulatory, structural, and assembly proteins of adherence fimbriae/pili, such as the P and K88 pili. These five genes and two adjacent ones which were not markedly related to proteins in the data bases comprise the pef (plasmid-encoded fimbriae) locus. Transposon TnphoA insertions in four genes in the pef locus (pefA, pefC, orf5 and orf6) resulted in active PhoA fusions and blocked or reduced the surface presentation of fimbriae, indicating that the proteins encoded by these four genes are translocated at least across the cytoplasmic membrane and contribute to formation of the fimbrial structure. The differences in genetic organization and protein sequence relatedness from other fimbrial gene clusters suggest that the pef locus might encode a novel type of fimbria. Between the pef and the repB loci, there were five open reading frames, one of which (orf8) gave rise to active PhoA fusions but was not necessary for fimbrial expression. Two of the other proteins were homologous to transcription regulatory proteins and a third was the rck gene, which encodes an outer membrane protein that confers complement resistance to serum-sensitive hosts.

Amino Acid Sequence↗

PubMed1990

Genes on the 90-kilobase plasmid of Salmonella typhimurium confer low-affinity cobalamin transport: relationship to fimbria biosynthesis genes.

A cloned fragment of Salmonella typhimurium DNA complemented the defect in cobalamin uptake of Escherichia coli or S. typhimurium btuB mutants, which lack the outer membrane high-affinity transport protein. This DNA fragment did not carry btuB and was derived from the 90-kb plasmid resident in S. typhimurium strains. The cobalamin transport activity engendered by this plasmid had substantially lower affinity and activity than that conferred by btuB. Complementation behavior and maxicell analyses of transposon insertions showed that the cloned fragment encoded five polypeptides, at least two of which were required for complementation activity. The nucleotide sequence of the coding region for one of these polypeptides, an outer membrane protein of about 84,000 Da, was determined. The deduced polypeptide had properties typical of outer membrane proteins, with an N-terminal signal sequence and a predicted preponderance of beta structure. This outer membrane protein had extensive amino acid sequence homology with PapC and FaeD, two E. coli outer membrane proteins involved in the export and assembly of pilus and fimbria subunits on the cell surface. This homology raises the likelihood that the observed cobalamin transport did not result from the production of an authentic transport system but that overexpression of one or more outer membrane proteins allowed leakage of cobalamins through the perturbed outer membrane. These results also suggest that the 90-kb plasmid carries genes encoding an adherence mechanism.

Amino Acid Sequence↗

PubMed1989

Nucleotide sequence of the Escherichia coli porin thermoregulatory gene envY.

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Amino Acid Sequence↗

PubMed1987

Nucleotide sequence of the uhp region of Escherichia coli.

The Escherichia coli uhp region encodes the transport system that mediates the uptake of a number of sugar phosphates as well as the regulatory components that are responsible for induction of this transport system by external glucose 6-phosphate. Four uhp genes have been identified by analysis of the complementation behavior and polypeptide coding capacity of plasmids carrying subcloned regions or transposon insertions. The nucleotide sequence of a 6.5-kilobase segment that contains the 3' end of the ilvBN operon and the entire uhp region was determined. Four open reading frames were identified in the locations expected for the various uhp genes; all were oriented in the same direction, counterclockwise relative to the genetic map. The properties of the polypeptides predicted from the nucleotide sequence were consistent with their observed features. The 196-amino-acid UhpA polypeptide has the composition characteristic of a soluble protein and bears homology to the DNA-binding regions of many regulatory activators and repressors. The 518-amino-acid UhpB and the 199-amino-acid UhpC regulatory proteins contain substantial segments of hydrophobic character. Similarly, the 463-amino-acid UhpT transporter is a hydrophobic protein with numerous potential transmembrane segments. The UhpC regulatory protein has substantial sequence homology to part of UhpT, suggesting that this regulatory protein might have evolved by duplication of the gene for the transporter and that its role in transmembrane signaling may involve sugar-phosphate-binding sites and transmembrane orientations similar to those of the transport protein.

Amino Acid Sequence↗

PubMed1986

Nucleotide sequence of the btuCED genes involved in vitamin B12 transport in Escherichia coli and homology with components of periplasmic-binding-protein-dependent transport systems.

The products of the btuCED region of the Escherichia coli chromosome participate in the transport of vitamin B12 across the cytoplasmic membrane. The nucleotide sequence of the 3,410-base-pair HindIII-HincII DNA fragment carrying a portion of the himA gene and the entire btuCED region was determined. Comparison of the location of the open reading frames with the gene boundaries defined by transposon insertions allowed the assignment of polypeptide products to gene sequences. The btuC product is a highly nonpolar integral membrane protein of molecular weight 31,683. The distribution of hydrophobic regions suggests the presence of numerous membrane-spanning domains. The btuD product is a relatively polar but membrane-associated polypeptide of Mr 27,088 and contains segments bearing extensive homology to the ATP-binding peripheral membrane constituents of periplasmic binding protein-dependent transport systems. Other regions of this protein are similar to portions of the outer membrane vitamin B12 receptor. The btuE product (Mr 20,474) appears to have a periplasmic location. It has the mean hydropathy of a soluble protein but lacks an obvious signal sequence. The cellular locations and structural and sequence homologies of the Btu polypeptides point to the similarity of these three proteins to components of binding protein-dependent transport systems. However, the dependence on a periplasmic vitamin B12-binding protein has not yet been demonstrated.

ATP-Binding Cassette Transporters↗

PubMedDate not supplied