scholarly journals Molecular and Immunological Analyses of the Mycobacterium avium Homolog of the Immunodominant Mycobacterium leprae 35-Kilodalton Protein

1998 ◽  
Vol 66 (6) ◽  
pp. 2684-2690 ◽  
Author(s):  
James A. Triccas ◽  
Nathalie Winter ◽  
Paul W. Roche ◽  
Andrea Gilpin ◽  
Kathleen E. Kendrick ◽  
...  
Keyword(s):  
Amino Acid ◽  
Mycobacterium Avium ◽  
Amino Acid Level ◽  
Streptomyces Griseus ◽  
Amino Acid Identity ◽  
Delayed Type Hypersensitivity ◽  
Gene Encoding ◽  
Genes Encoding ◽  
Species Specific ◽  
35 Kda Protein

ABSTRACT The analysis of host immunity to mycobacteria and the development of discriminatory diagnostic reagents relies on the characterization of conserved and species-specific mycobacterial antigens. In this report, we have characterized the Mycobacterium avium homolog of the highly immunogenic M. leprae 35-kDa protein. The genes encoding these two proteins were well conserved, having 82% DNA identity and 90% identity at the amino acid level. Moreover both proteins, purified from the fast-growing host M. smegmatis, formed multimeric complexes of around 1000 kDa in size and were antigenically related as assessed through their recognition by antibodies and T cells from M. leprae-infected individuals. The 35-kDa protein exhibited significant sequence identity with proteins from Streptomyces griseus and the cyanobacterium Synechoccocus sp. strain PCC 7942 that are up-regulated under conditions of nutrient deprivation. The 67% amino acid identity between the M. avium 35-kDa protein and SrpI of Synechoccocus was spread across the sequences of both proteins, while the homologous regions of the 35-kDa protein and the P3 sporulation protein of S. griseus were interrupted in the P3 protein by a divergent central region. Assessment by PCR demonstrated that the gene encoding the M. avium35-kDa protein was present in all 30 M. avium clinical isolates tested but absent from M. intracellulare,M. tuberculosis, or M. bovis BCG. Mice infected with M. avium, but not M. bovis BCG, developed specific immunoglobulin G antibodies to the 35-kDa protein, consistent with the observation that tuberculosis patients do not recognize the antigen. Strong delayed-type hypersensitivity was elicited by the protein in guinea pigs sensitized with M. avium.

scholarly journals A50 Whole-genome sequencing of African swine fever isolates from Sardinia

2019 ◽  
Vol 5 (Supplement_1) ◽  
Author(s):  
C Torresi ◽  
F Granberg ◽  
L Bertolotti ◽  
A Oggiano ◽  
B Colitti ◽  
...  
Keyword(s):  
Amino Acid ◽  
Sequence Data ◽  
Temporal Distribution ◽  
African Swine Fever ◽  
Amino Acid Identity ◽  
Genotype I ◽  
Acid Identity ◽  
Wide Range ◽  
Intergenic Sequences ◽  
Genes Encoding

Abstract In order to assess the molecular epidemiology of African swine fever (ASF) in Sardinia, we analyzed a wide range of isolates from wild and domestic pigs over a 31-year period (1978–2009) by genotyping sequence data from the genes encoding the p54 and the p72 proteins and the CVR. On this basis, the analysis of the B602L gene revealed a minor difference, placing the Sardinian isolates into two clusters according to their temporal distribution. As an extension of this study, in order to achieve a higher level of discrimination, three further variable genome regions, namely p30, CD2v, and I73R/I329L, of a large number of isolates collected from outbreaks in the years 2002–14 have been investigated. Sequence analysis of the CD2v region revealed a temporal subdivision of the viruses into two subgroups. These data, together with those from the B602L gene analysis, demonstrated that the viruses circulating in Sardinia belong to p72/genotype I, but since 1990 have undergone minor genetic variations in respect to its ancestor, thus making it impossible to trace isolates, enabling a more accurate assessment of the origin of outbreaks, and extending knowledge of virus evolution. To solve this problem, we have sequenced and annotated the complete genome of nine ASF isolates collected in Sardinia between 1978 and 2012. This was achieved using sequence data determined by next-generation sequencing. The results showed a very high identity with range of nucleotide similarity among isolates of 99.5 per cent to 99.9 per cent. The ASF virus (ASFV) genomes were composed of terminal inverted repeats and conserved and non-conserved ORFs. Among the conserved ORFs, B385R, H339R, and O61R-p12 showed 100 per cent amino acid identity. The same was true for the hypervariable ORFs, with regard to X69R, DP96R, DP60R, EP153R, B407L, I10L, and L60L genes. The EP402R and B602L genes showed, as expected, an amino acid identity range of 98.5 per cent to 100 per cent and 91 per cent to 100 per cent, respectively. In addition, all of the isolates displayed variable intergenic sequences. As a whole, the results from our studies confirmed a remarkable genetic stability of the ASFV/p72 genotype I viruses circulating in Sardinia.

The gene encoding a major component of the lateral elements of synaptonemal complexes of the rat is related to X-linked lymphocyte-regulated genes

1994 ◽  
Vol 14 (2) ◽  
pp. 1137-1146
Author(s):  
J H Lammers ◽  
H H Offenberg ◽  
M van Aalderen ◽  
A C Vink ◽  
A J Dietrich ◽  
...  
Keyword(s):  
Amino Acid ◽  
Sequence Similarity ◽  
Sodium Dodecyl ◽  
Coiled Coil ◽  
Amino Acid Identity ◽  
Male Rat ◽  
Amino Acid Sequence Similarity ◽  
Gene Encoding ◽  
Synaptonemal Complexes ◽  
Complex Protein

The lateral elements of synaptonemal complexes (SCs) of the rat contain major components with relative electrophoretic mobilities (M(r)S) of 30,000 and 33,000. After one-dimensional separation of SC proteins on polyacrylamide-sodium dodecyl sulfate gels, these components show up as two broad bands. These bands contain closely related proteins, as judged from their peptide maps and immunological reactivity. Using affinity-purified polyclonal anti-30,000- and anti-33,000-M(r) component antibodies, we isolated a cDNA encoding at least one of the 30,000- or 33,000-M(r) SC components. The protein predicted from the nucleotide sequence of the cDNA, called SCP3 (for synaptonemal complex protein 3), has a molecular mass of 29.7 kDa and a pI value of 9.4. It has a potential nucleotide binding site and contains stretches that are predicted to be capable of forming coiled-coil structures. In the male rat, the gene encoding SCP3 is transcribed exclusively in the testis. SCP3 has significant amino acid similarity to the pM1 protein, which is one of the predicted products of an X-linked lymphocyte-regulated gene family of the mouse: there are 63% amino acid sequence similarity and 35% amino acid identity between the SCP3 and pM1 proteins. However, SCP3 differs from pM1 in several respects, and whether the proteins fulfill related functions is still an open question.

scholarly journals Rho1p-Bni1p-Spa2p Interactions: Implication in Localization of Bni1p at the Bud Site and Regulation of the Actin Cytoskeleton inSaccharomyces cerevisiae

1998 ◽  
Vol 9 (5) ◽  
pp. 1221-1233 ◽  
Author(s):  
Takeshi Fujiwara ◽  
Kazuma Tanaka ◽  
Akihisa Mino ◽  
Mitsuhiro Kikyo ◽  
Kazuo Takahashi ◽  
...  
Keyword(s):  
Amino Acid ◽  
Actin Cytoskeleton ◽  
Binding Protein ◽  
Microscopic Analysis ◽  
Mitogen Activated Protein Kinase ◽  
Gene Encoding ◽  
Amino Acid Region ◽  
Genes Encoding ◽  
Mitogen Activated Protein ◽  
Acid Region

Rho1p is a yeast homolog of mammalian RhoA small GTP-binding protein. Rho1p is localized at the growth sites and required for bud formation. We have recently shown that Bni1p is a potential target of Rho1p and that Bni1p regulates reorganization of the actin cytoskeleton through interactions with profilin, an actin monomer-binding protein. Using the yeast two-hybrid screening system, we cloned a gene encoding a protein that interacted with Bni1p. This protein, Spa2p, was known to be localized at the bud tip and to be implicated in the establishment of cell polarity. The C-terminal 254 amino acid region of Spa2p, Spa2p(1213–1466), directly bound to a 162-amino acid region of Bni1p, Bni1p(826–987). Genetic analyses revealed that both thebni1 and spa2 mutations showed synthetic lethal interactions with mutations in the genes encoding components of the Pkc1p-mitogen-activated protein kinase pathway, in which Pkc1p is another target of Rho1p. Immunofluorescence microscopic analysis showed that Bni1p was localized at the bud tip in wild-type cells. However, in the spa2 mutant, Bni1p was not localized at the bud tip and instead localized diffusely in the cytoplasm. A mutant Bni1p, which lacked the Rho1p-binding region, also failed to be localized at the bud tip. These results indicate that both Rho1p and Spa2p are involved in the localization of Bni1p at the growth sites where Rho1p regulates reorganization of the actin cytoskeleton through Bni1p.

The Mycobacterium avium subsp. paratuberculosis 35 kDa protein plays a role in invasion of bovine epithelial cells

Microbiology ◽  
2003 ◽  
Vol 149 (8) ◽  
pp. 2061-2069 ◽  
Author(s):  
John P. Bannantine ◽  
Jason F. J. Huntley ◽  
Elizabeth Miltner ◽  
Judith R. Stabel ◽  
Luiz E. Bermudez
Keyword(s):  
Epithelial Cells ◽  
Mycobacterium Avium ◽  
Immunoblot Analysis ◽  
Kidney Cells ◽  
Intestinal Epithelial ◽  
Low Oxygen ◽  
Gene Encoding ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
35 Kda Protein

Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis) enters intestinal epithelial cells of cattle and other ruminants via a mechanism that remains to be fully elucidated. This study showed that a gene encoding the M. paratuberculosis 35 kDa major membrane protein (MMP) is expressed at a higher level in low-oxygen and high-osmolarity conditions that are similar to the environment of the intestine. In addition, cattle with Johne's disease produced antibodies against MMP, suggesting that the protein is present during infection. The gene encoding MMP was cloned and expressed as a fusion protein with the maltose-binding protein (MBP–MMP) in Escherichia coli. Rabbit antisera were raised against a M. paratuberculosis whole-cell sonicate and MMP-specific antibodies were purified from these sera by affinity chromatography. MMP was localized to the surface of M. paratuberculosis by immunoelectron microscopy and by immunoblot analysis of fractionated protein lysates. Both anti-MMP antibodies and MBP–MMP protein inhibited M. paratuberculosis invasion of cultured Madin–Darby bovine kidney cells by 30 %. In similar invasion experiments with M. paratuberculosis incubated in low oxygen tension, these antibodies and protein decreased invasion by 60 %. Collectively, these data show that the 35 kDa MMP is a surface exposed protein that plays a role in invasion of epithelial cells. The authors suggest that the MMP is a virulence factor of M. paratuberculosis that may be important in the initiation of infection in vivo.

scholarly journals Staphylococcus caprae Strains Carry Determinants Known To Be Involved in Pathogenicity: a Gene Encoding an Autolysin-Binding Fibronectin and the ica Operon Involved in Biofilm Formation

2001 ◽  
Vol 69 (2) ◽  
pp. 712-718 ◽  
Author(s):  
Jeanine Allignet ◽  
Sylvie Aubert ◽  
Keith G. H. Dyke ◽  
Nevine El Solh
Keyword(s):  
Amino Acid ◽  
Biofilm Formation ◽  
Insertion Sequence ◽  
Mobile Element ◽  
Amino Acid Identity ◽  
Gene Encoding ◽  
Surface Anchoring ◽  
Acid Identity ◽  
Putative Signal Peptide ◽  
Fibronectin Binding

ABSTRACT The atlC gene (1,485 bp), encoding an autolysin which binds fibronectin, and the ica operon, involved in biofilm formation, were isolated from the chromosome of an infectious isolate of Staphylococcus caprae and sequenced. AtlC (155 kDa) is similar to the staphylococcal autolysins Atl, AtlE, Aas (48 to 72% amino acid identity) and contains a putative signal peptide of 29 amino acids and two enzymatic centers (N-acetylmuramoyl-l-alanine amidase and endo-β-N-acetylglucosaminidase) interconnected by three imperfect fibronectin-binding repeats. The glycine-tryptophan (GW) motif found in the central and end part of each repeat may serve for cell surface anchoring of AtlC as they do in Listeria monocytogenes. The S. caprae ica operon contains four genes closely related to S. epidermidis and S. aureus icaA, icaB, icaC, and icaDgenes (≥ 68% similarity) and is preceded by a gene similar toicaR (≥70% similarity). The polypeptides deduced from theS. caprae ica genes exhibit 67 to 88% amino acid identity to those of S. epidermidis and S. aureus icagenes. The ica operon and icaR gene were analyzed in 14 S. caprae strains from human specimens or goats' milk. Some of the strains produced biofilm, and others did not. All strains carry the ica operon and icaR of the same sizes and in the same relative positions, suggesting that the absence of biofilm formation is not related to the insertion of a mobile element such as an insertion sequence or a transposon.

scholarly journals Sulfonamide Resistance in Streptococcus pyogenes Is Associated with Differences in the Amino Acid Sequence of Its Chromosomal Dihydropteroate Synthase

1998 ◽  
Vol 42 (5) ◽  
pp. 1062-1067 ◽  
Author(s):  
Göte Swedberg ◽  
Signe Ringertz ◽  
Ola Sköld
Keyword(s):  
Amino Acid ◽  
Streptococcus Pyogenes ◽  
Gtp Cyclohydrolase ◽  
Gene Encoding ◽  
Additional Indication ◽  
Kinetic Measurements ◽  
Dihydropteroate Synthase ◽  
Development Of Resistance ◽  
Sulfonamide Resistance ◽  
Genes Encoding

ABSTRACT Sulfonamide resistance in recent isolates of Streptococcus pyogenes was found to be associated with alterations of the chromosomally encoded dihydropteroate synthase (DHPS). There were 111 different nucleotides (13.8%) in the genes found in susceptible and resistant isolates, respectively, resulting in 30 amino acid changes (11.3%). These substantial changes suggested the possibility of a foreign origin of the resistance gene, in parallel to what has already been found for sulfonamide resistance in Neisseria meningitidis. The gene encoding DHPS was linked to at least three other genes encoding enzymes of the folate pathway. These genes were in the order GTP cyclohydrolase, dihydropteroate synthase, dihydroneopterin aldolase, and hydroxymethyldihydropterin pyrophosphokinase. The nucleotide differences in genes from resistant and susceptible strains extended from the beginning of the GTP cyclohydrolase gene to the end of the gene encoding DHPS, an additional indication for gene transfer in the development of resistance. Kinetic measurements established different affinities for sulfathiazole for DHPS enzymes isolated from resistant and susceptible strains.

scholarly journals Biochemical and Genetic Characterization of the vanC-2 Vancomycin Resistance Gene Cluster of Enterococcus casseliflavus ATCC 25788

2002 ◽  
Vol 46 (10) ◽  
pp. 3125-3132 ◽  
Author(s):  
Ireena Dutta ◽  
Peter E. Reynolds
Keyword(s):  
Amino Acid ◽  
Gene Cluster ◽  
Response Regulator ◽  
Extended Period ◽  
Amino Acid Identity ◽  
Serine Racemase ◽  
Gene Encoding ◽  
Acid Identity ◽  
Induction Process ◽  
Enterococcus Casseliflavus

ABSTRACT The vanC-2 cluster of Enterococcus casseliflavus ATCC 25788 consisted of five genes (vanC-2, vanXYC-2 , vanTC-2 , vanRC-2 , and vanSC-2 ) and shared the same organization as the vanC cluster of E. gallinarum BM4174. The proteins encoded by these genes displayed a high degree of amino acid identity to the proteins encoded within the vanC gene cluster. The putative d,d-dipeptidase-d,d-carboxypeptidase, VanXYC-2, exhibited 81% amino acid identity to VanXYC, and VanTC-2 displayed 65% amino acid identity to the serine racemase, VanT. VanRC-2 and VanSC-2 displayed high degrees of identity to VanRC and VanSC, respectively, and contained the conserved residues identified as important to their function as a response regulator and histidine kinase, respectively. Resistance to vancomycin was expressed inducibly in E. casseliflavus ATCC 25788 and required an extended period of induction. Analysis of peptidoglycan precursors revealed that UDP-N-acetylmuramyl-l-Ala-δ-d-Glu-l-Lys-d-Ala-d-Ser could not be detected until several hours after the addition of vancomycin, and its appearance coincided with the resumption of growth. The introduction of additional copies of the vanTC-2 gene, encoding a putative serine racemase, and the presence of supplementary d-serine in the growth medium both significantly reduced the period before growth resumed after addition of vancomycin. This suggested that the availability of d-serine plays an important role in the induction process.

scholarly journals Purification, Characterization, and Molecular Analysis of the Gene Encoding Glucosyltransferase fromStreptococcus oralis

2000 ◽  
Vol 68 (5) ◽  
pp. 2475-2483 ◽  
Author(s):  
Taku Fujiwara ◽  
Tomonori Hoshino ◽  
Takashi Ooshima ◽  
Shizuo Sobue ◽  
Shigeyuki Hamada
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Pcr Primers ◽  
Water Soluble ◽  
Oral Streptococci ◽  
Amino Acid Residues ◽  
Gene Encoding ◽  
Streptococcus Sanguis ◽  
Specific Nucleotide Sequence ◽  
Species Specific

ABSTRACT Streptococcus oralis is a member of the oral streptococcal family and an early-colonizing microorganism in the oral cavity of humans. S. oralis is known to produce glucosyltransferase (GTase), which synthesizes glucans from sucrose. The enzyme was purified chromatographically from a culture supernatant of S. oralis ATCC 10557. The purified enzyme, GTase-R, had a molecular mass of 173 kDa and a pI of 6.3. This enzyme mainly synthesized water-soluble glucans with no primer dependency. The addition of GTase markedly enhanced the sucrose-dependent resting cell adhesion of Streptococcus mutans at a level similar to that found in growing cells of S. mutans. The antibody against GTase-R inhibited the glucan-synthesizing activities ofStreptococcus gordonii and Streptococcus sanguis, as well as S. oralis. The N-terminal amino acid sequence of GTase-R exhibited no similarities to known GTase sequences of oral streptococci. Using degenerate PCR primers, an 8.1-kb DNA fragment, carrying the gene (gtfR) coding for GTase-R and its regulator gene (rgg), was cloned and sequenced. Comparison of the deduced amino acid sequence revealed that thergg genes of S. oralis and S. gordonii exhibited a close similarity. The gtfR gene was found to possess a species-specific nucleotide sequence corresponding to the N-terminal 130 amino acid residues. Insertion oferm or aphA into the rgg orgtfR gene resulted in decreased GTase activity by the organism and changed the colony morphology of these transformants. These results indicate that S. oralis GTase may play an important role in the subsequent colonizing of mutans streptoccoci.

scholarly journals Reconstructing Genomes of Carbon Monoxide Oxidisers in Volcanic Deposits Including Members of the Class Ktedonobacteria

2020 ◽  
Vol 8 (12) ◽  
pp. 1880
Author(s):  
Marcela Hernández ◽  
Blanca Vera-Gargallo ◽  
Marcela Calabi-Floody ◽  
Gary M. King ◽  
Ralf Conrad ◽  
...  
Keyword(s):  
Carbon Monoxide ◽  
Carbon Fixation ◽  
Large Subunit ◽  
Natural Habitat ◽  
Volcanic Rocks ◽  
Amino Acid Identity ◽  
Cellular Functions ◽  
Gene Encoding ◽  
Genes Encoding ◽  
Successional Stages

Microorganisms can potentially colonise volcanic rocks using the chemical energy in reduced gases such as methane, hydrogen (H2) and carbon monoxide (CO). In this study, we analysed soil metagenomes from Chilean volcanic soils, representing three different successional stages with ages of 380, 269 and 63 years, respectively. A total of 19 metagenome-assembled genomes (MAGs) were retrieved from all stages with a higher number observed in the youngest soil (1640: 2 MAGs, 1751: 1 MAG, 1957: 16 MAGs). Genomic similarity indices showed that several MAGs had amino-acid identity (AAI) values >50% to the phyla Actinobacteria, Acidobacteria, Gemmatimonadetes, Proteobacteria and Chloroflexi. Three MAGs from the youngest site (1957) belonged to the class Ktedonobacteria (Chloroflexi). Complete cellular functions of all the MAGs were characterised, including carbon fixation, terpenoid backbone biosynthesis, formate oxidation and CO oxidation. All 19 environmental genomes contained at least one gene encoding a putative carbon monoxide dehydrogenase (CODH). Three MAGs had form I coxL operon (encoding the large subunit CO-dehydrogenase). One of these MAGs (MAG-1957-2.1, Ktedonobacterales) was highly abundant in the youngest soil. MAG-1957-2.1 also contained genes encoding a [NiFe]-hydrogenase and hyp genes encoding accessory enzymes and proteins. Little is known about the Ktedonobacterales through cultivated isolates, but some species can utilise H2 and CO for growth. Our results strongly suggest that the remote volcanic sites in Chile represent a natural habitat for Ktedonobacteria and they may use reduced gases for growth.

scholarly journals Characterization of Staphylococcus pseudintermedius isolated from diseased dogs in Lithuania

2016 ◽  
Vol 19 (1) ◽  
pp. 7-14 ◽  
Author(s):  
M. Ruzauskas ◽  
N. Couto ◽  
A. Pavilonis ◽  
I. Klimiene ◽  
R. Siugzdiniene ◽  
...  
Keyword(s):  
Companion Animals ◽  
Penicillin G ◽  
Special Focus ◽  
Reproductive Disorders ◽  
Gene Encoding ◽  
Staphylococcus Pseudintermedius ◽  
Genes Encoding ◽  
Species Specific ◽  
Staphylococcus Spp

AbstractThe aim of this study was to characterize Staphylococcus pseudintermedius for its antimicrobial resistance and virulence factors with a special focus on methicillin-resistant (MRSP) strains isolated from sick dogs in Lithuania. Clinically sick adult dogs suffering from infections (n=214) and bitches with reproductive disorders (n=36) from kennels were selected for the study. Samples (n=192) from the 250 tested (76.8%) dogs were positive for Staphylococcus spp. Molecular profiling using the species-specific nuc gene identified 51 isolates as S. pseudintermedius (26.6% from a total number of isolated staphylococci) of which 15 isolates were identified as MRSP. Ten MRSP isolates were isolated from bitches with reproductive disorders from two large breeding kennels. Data on susceptibility of S. pseudintermedius to different antimicrobials revealed that all isolates were susceptible to vancomycin, daptomycin and linezolid. Two isolates (3.9%) were resistant to rifampicin. A high resistance was seen towards penicillin G (94.1%), tetracycline (64.7%) and macrolides (68.7%). Resistance to fluoroquinolones ranged from 25.5% (gatifloxacin) to 31.4% (ciprofloxacin). The most prevalent genes encoding resistance included blaZ, aac(6’)-Ie-aph(2’’)-Ia, mecA, and tet(M). The Luk-I gene encoding a leukotoxin was detected in 29% of the isolates, whereas the siet gene encoding exfoliative toxin was detected in 69% of the S. pseudintermedius isolates. This report of MRSP in companion animals represents a major challenge for veterinarians in terms of antibiotic therapy and is a concern for both animal and public health.

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