Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0023241 (Legionella)
 6,990 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The enzyme activities of four strains of Legionella pneumophilia were investigated by using the API ZYM system (API System S.A., F-38390 Montalieu Vercieu, France) and synthetic substrates. Aminopeptidases were detected specifically against L-alanine, L-arginine, L-aspartic acid, L-cystine, L-glutaminic acid, glycine, L-histidine, L-isoleucine, L-leucine, L-lysine, L-methionine, L-phenylalanine, L-tryptophan, L-tyrosine, and L-valine. Furthermore, the bacteria possesses esterase activity splitting propionate, butyrate, caproate, caprylate, and caprate, but not laurate, myristate, palmitate, and stearate, esters. The enzymes studies were inhibited partially by aprotinin. No inhibition of phosphatase (pH range, 5.4 to 8.5) or of phosphoamidase was observed. Activities of arylsulfatase, chymotrypsin, trypsin, and glycosidases could not be detected.
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PMID:Enzymatic profile of Legionella pneumophilia. 616 35

The culture conditions of Afipia felis, A. broomeae, A. clevelandensis and three unnamed Afipia genospecies were investigated on BCY agar supplemented with different substances known as growth factors of Legionella spp. and, furthermore, with sodium chloride and other salts. The organisms were found to be susceptible to a certain degree to byproducts of the autoclaving which are scavenged by activated by charcoal. Growth was weakly enhanced by ferric pyrophosphate, cystein.HCl, and alpha-ketoglutarate. These substances are no obligatory growth factors. The optimal pH value was about 6.8. Afipia spp. showed a strong susceptibility to NaCl and other salts. They possess phosphatase, phosphoamidase, phosphodiesterase, a weak sulfatase, glycine aminopeptidase, and L-lysine aminopeptidase. The strains differed with regard to other proteases and aminopeptidases. The decimal reduction times of A. felis at 55 degrees C and 60 degrees C were 11 min, < 1 min, respectively.
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PMID:Investigations of culture and properties of Afipia spp. 773 25

The bacterium Legionella pneumophila is the responsible agent for Legionnaires' disease and has recently been shown to harbor a gene encoding a kinase that confers resistance to the aminoglycoside antibiotic spectinomycin (Suter, T. M., Viswanathan, V. K., and Cianciotto, N. P. (1997) Antimicrob. Agents Chemother. 41, 1385-1388). We report the overproduction, purification, and characterization of this spectinomycin kinase from an expressing system in Escherichia coli. The purified protein shows stringent substrate specificity for spectinomycin with Km = 21.5 microM and kcat = 24.2 s-1 and does not bind other aminoglycosides including kanamycin, amikacin, neomycin, butirosin, streptomycin, or apramycin. Purification of spectinomycin phosphate followed by characterization by mass spectrometry and 1H, 13C, and 31P NMR established the site of phosphorylation to be at the hydroxyl group at position 9. Thus this enzyme is designated APH(9)-Ia (where APH is aminoglycoside kinase). The enzyme was inactivated by the electrophilic ATP analogue 5'-[p-(fluorosulfonyl)benzoyl]adenosine, consistent with a nucleophilic residue such as Lys lining the nucleotide binding pocket. Site-directed mutagenesis of Lys-52 and Asp-212 to Ala confirmed that these residues were important for catalysis, with Lys-52 playing a potential role in ATP binding and Asp-212 in phosphoryl transfer. Thio and solvent isotope effect experiments in the presence of either Mg2+ or Mn2+ were consistent with a kinetic mechanism in which phosphate transfer does not contribute significantly to the rate-limiting step. These results establish that APH(9)-Ia is a highly specific antibiotic resistance kinase and provides the requisite mechanistic information for future structural studies.
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PMID:Spectinomycin kinase from Legionella pneumophila. Characterization of substrate specificity and identification of catalytically important residues. 961 79

The potential for selection in vitro of Legionella pneumophila mutants resistant to fluoroquinolones was investigated. Six distinct clinical isolates of L. pneumophila were subcultured in subinhibitory concentrations of ciprofloxacin, levofloxacin, clinafloxacin, trovafloxacin and moxifloxacin until MICs increased at least eight-fold. The numbers of serial passages required in microbroth dilution series were determined. The gyrA gene of the six parental strains, and 12 selected mutant strains, was sequenced. The five quinolones differed markedly in their ability to select mutants with decreased susceptibility. The average number of serial passages required was low in the cases of clinafloxacin (n = 10.6), ciprofloxacin and levofloxacin (both n = 13), but notably higher for trovafloxacin (n = 26.6) and moxifloxacin (n = 22.5). Five mutants treated with ciprofloxacin and three treated with moxifloxacin showed Thr83-->Lys or Thr83-->Ile amino acid changes in the gyrA gene. In conclusion, different quinolones lose their antimicrobial effect after a varying number of passages. This study demonstrated, for the first time to our knowledge, that gyrA in L. pneumophila is a possible target of fluoroquinolones.
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PMID:Development and mechanism of fluoroquinolone resistance in Legionella pneumophila. 1256 91

The ability of Legionella pneumophila to grow and cause disease in the host is completely dependent on a type IV secretion system known as the Dot/Icm complex. This membrane-spanning apparatus translocates effector molecules into host cells in a process that is poorly understood but that is known to require the putative ATPase DotB. One possible role for DotB is suggested by its similarity to the PilT family of proteins, which mediate pilus retraction. To better understand the molecular behavior of DotB, we have purified the protein and shown that it forms stable homohexameric rings and hydrolyzes ATP with a specific activity of 6.4 nmol of ATP/min/mg of protein. ATPase activity is critical to the function of DotB, as alteration of the conserved Walker box lysine residue resulted in a mutant protein, DotB K162Q, which failed to bind or hydrolyze ATP and which could not complement a DeltadotB strain for intracellular growth in macrophages. Consistent with the ability of DotB to interact with itself, the dotBK162Q allele exhibited transdominance over wild-type dotB, providing the first example of such a mutation in L. pneumophila. Finally, the DotB K162Q mutant protein had a significantly enhanced membrane localization in L. pneumophila compared to wild-type DotB, suggesting a relationship between nucleotide binding and membrane association. These results are consistent with a model in which DotB cycles between the cytoplasm and the Dot/Icm complex at the membrane, where it hydrolyzes nucleotides to provide energy to the complex.
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PMID:The Legionella pneumophila PilT homologue DotB exhibits ATPase activity that is critical for intracellular growth. 1499 96

Nearly two dozen microbial pathogens have surface polysaccharides or lipo-oligosaccharides that contain sialic acid (Sia), and several Sia-dependent virulence mechanisms are known to enhance bacterial survival or result in host tissue injury. Some pathogens are also known to O-acetylate their Sias, although the role of this modification in pathogenesis remains unclear. We report that neuD, a gene located within the Group B Streptococcus (GBS) Sia biosynthetic gene cluster, encodes a Sia O-acetyltransferase that is itself required for capsular polysaccharide (CPS) sialylation. Homology modeling and site-directed mutagenesis identified Lys-123 as a critical residue for Sia O-acetyltransferase activity. Moreover, a single nucleotide polymorphism in neuD can determine whether GBS displays a "high" or "low" Sia O-acetylation phenotype. Complementation analysis revealed that Escherichia coli K1 NeuD also functions as a Sia O-acetyltransferase in GBS. In fact, NeuD homologs are commonly found within Sia biosynthetic gene clusters. A bioinformatic approach identified 18 bacterial species with a Sia biosynthetic gene cluster that included neuD. Included in this list are the sialylated human pathogens Legionella pneumophila, Vibrio parahemeolyticus, Pseudomonas aeruginosa, and Campylobacter jejuni, as well as an additional 12 bacterial species never before analyzed for Sia expression. Phylogenetic analysis shows that NeuD homologs of sialylated pathogens share a common evolutionary lineage distinct from the poly-Sia O-acetyltransferase of E. coli K1. These studies define a molecular genetic approach for the selective elimination of GBS Sia O-acetylation without concurrent loss of sialylation, a key to further studies addressing the role(s) of this modification in bacterial virulence.
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PMID:The group B streptococcal sialic acid O-acetyltransferase is encoded by neuD, a conserved component of bacterial sialic acid biosynthetic gene clusters. 1649 Jul 81

Legionella pneumophila is an intracellular pathogen that replicates in a unique vacuole that avoids endocytic maturation. Previous studies have shown host vesicles attached to the L. pneumophila-containing vacuole (LCV) minutes after uptake. Here we examine the origin and content of these vesicles by electron microscopy (EM). Our data demonstrate that the attached vesicles are derived from endoplasmic reticulum (ER) based the presence of the resident ER proteins glucose-6-phosphatase, protein disulphide isomerase (PDI) and proteins having the ER-retention signal lysine-aspartic acid-glutamic acid-leucine (KDEL). After tethering occurred, ER markers inside of attached vesicles were delivered into the lumen of the LCV, indicating ER fusion. Treatment of cells with brefeldin A did not interfere with the attachment of ER vesicles with the LCV, suggesting that tethering of these vesicles does not require activities mediated by ADP-ribosylation factor (ARF). ER vesicles were not tethered to the LCV in cells producing the Sar1H79G protein, indicating that vesicles produced by the Sar1/CopII system are necessary for vesicle attachment. From these data we conclude that formation of the organelle that supports L. pneumophila replication is a two-stage process that involves remodelling of the LCV by early secretory vesicles produced by the Sar1/CopII system, followed by attachment and fusion of ER.
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PMID:Attachment and fusion of endoplasmic reticulum with vacuoles containing Legionella pneumophila. 1661 Dec 28

Legionella pneumophila, the agent of Legionnaires' disease, is an intracellular parasite of aquatic amoebae and human macrophages. A key factor for L. pneumophila in intracellular infection is its type II protein secretion system (Lsp). In order to more completely define Lsp output, we recently performed a proteomic analysis of culture supernatants. Based upon the predictions of that analysis, we found that L. pneumophila secretes two distinct aminopeptidase activities encoded by the genes lapA and lapB. Whereas lapA conferred activity against leucine, phenylalanine, and tyrosine aminopeptides, lapB was linked to the cleavage of lysine- and arginine-containing substrates. To assess the role of secreted aminopeptidases in intracellular infection, we examined the relative abilities of lapA and lapB mutants to infect human U937 cell macrophages as well as Hartmannella vermiformis and Acanthamoeba castellanii amoebae. Although these experiments identified a dispensable role for LapA and LapB, they uncovered a previously unrecognized role for the type II-dependent ProA (MspA) metalloprotease. Whereas proA mutants were not defective for macrophage or A. castellanii infection, they (but not their complemented derivatives) were impaired for growth upon coculture with H. vermiformis. Thus, ProA represents the first type II effector implicated in an intracellular infection event. Furthermore, proA represents an L. pneumophila gene that shows differential importance among protozoan infection models, suggesting that the legionellae might have evolved some of its factors to especially target certain of their protozoan hosts.
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PMID:The type II secretion system of Legionella pneumophila elaborates two aminopeptidases, as well as a metalloprotease that contributes to differential infection among protozoan hosts. 1808 80

The opportunistic pathogen Legionella pneumophila replicates in human lung macrophages and in free-living amoebae. To accommodate the transfer between host cells, L. pneumophila switches from a replicative to a transmissive phase. L. pneumophila harbors a gene cluster homologous to the Vibrio cholerae cqsAS quorum sensing system, encoding a putative autoinducer synthase (lqsA) and a sensor kinase (lqsS), which flank a response regulator (lqsR). LqsR is an element of the L. pneumophila virulence regulatory network, which promotes pathogen-host cell interactions and inhibits entry into the replicative growth phase. Here, we show that lqsA functionally complements a V. cholerae cqsA autoinducer synthase deletion mutant and, upon expression in L. pneumophila or Escherichia coli, produces the diffusible signaling molecule LAI-1 (Legionella autoinducer-1). LAI-1 is distinct from CAI-1 (Cholerae autoinducer-1) and was identified as 3-hydroxypentadecan-4-one using liquid chromatography coupled to high resolution tandem mass spectrometry. The activity of both LqsA and CqsA was abolished upon mutation of a conserved lysine, and covalent binding of the cofactor pyridoxal 5'-phosphate to this lysine was confirmed by mass spectrometry. Thus, LqsA and CqsA belong to a family of pyridoxal 5'-phosphate-dependent autoinducer synthases, which produce the alpha-hydroxyketone signaling molecules LAI-1 and CAI-1.
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PMID:The Legionella autoinducer synthase LqsA produces an alpha-hydroxyketone signaling molecule. 1841 Dec 63

Legionella pneumophila causes severe pneumonia. Acetylation of histones is thought to be an important regulator of gene transcription, but its impact on L. pneumophila-induced expression of proinflammatory cytokines is unknown. L. pneumophila strain 130b induced the expression of the important chemoattractant IL-8 and genome-wide histone modifications in human lung epithelial A549 cells. We analyzed the IL-8-promoter and found that histone H4 was acetylated and H3 was phosphorylated at Ser(10) and acetylated at Lys(14), followed by transcription factor NF-kappaB. Recruitment of RNA polymerase II to the IL-8 promoter corresponded with increases in gene transcription. Histone modification and IL-8 release were dependent on p38 kinase and NF-kappaB pathways. Legionella-induced IL-8 expression was decreased by histone acetylase (HAT) inhibitor anacardic acid and enhanced by histone deacetylase (HDAC) inhibitor trichostatin A. After Legionella infection, HATs p300 and CREB-binding protein were time-dependently recruited to the IL-8 promoter, whereas HDAC1 and HDAC5 first decreased and later reappeared at the promoter. Legionella specifically induced expression of HDAC5 but not of other HDACs in lung epithelial cells, but knockdown of HDAC1 or 5 did not alter IL-8 release. Furthermore, Legionella-induced cytokine release, promoter-specific histone modifications, and RNA polymerase II recruitment were reduced in infection with flagellin-deletion mutants. Legionella-induced histone modification as well as HAT-/HDAC-dependent IL-8 release could also be shown in primary lung epithelial cells. In summary, histone acetylation seems to be important for the regulation of proinflammatory gene expression in L. pneumophila infected lung epithelial cells. These pathways may contribute to the host response in Legionnaires' disease.
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PMID:Histone acetylation and flagellin are essential for Legionella pneumophila-induced cytokine expression. 1860 45


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