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Biomedical subjects

G Christiansen

Publications and source records attributed to G Christiansen.

At least 73 records · Page 4Linked to original sources
PubMed1994

Analysis of a Mycoplasma hominis membrane protein, P120.

The monoclonal antibody mAb 26.7D generated against a clinical isolate of Mycoplasma hominis 7488 was shown to react with a surface-exposed epitope on a 120-kDa protein (P120). The gene encoding the protein was cloned and sequenced, and the transcriptional start point was determined by primer extension analysis. The gene contained an open reading frame of 3237 bp encoding a peptide of 1079 amino acids with a deduced molecular mass of 123 kDa. A putative amino-terminal signal peptide and cleavage site for signal peptidase II were found. This suggests that the protein was synthesized as a precursor with subsequent processing to a mature lipoprotein. Surface exposure was confirmed by immunoelectron microscopy. mAb 26.7D reacted with 11 of 19 M. hominis strains. The gene was, however, present in all strains.

Amino Acid Sequence↗
PubMed1994

Characterization of a Mycoplasma hominis gene encoding lysyl-tRNA synthetase (LysRS).

The gene encoding lysyl-tRNA synthetase (lysS) in Mycoplasma hominis was cloned and sequenced. The gene was found to have an open reading frame of 1466 bp encoding a polypeptide with a predicted molecular mass of 57 kDa. The amino acid sequence showed 44.3% and 43.7% identity to the Escherichia coli lysyl-tRNA synthetases, encoded by lysS and lysU. Only one lysyl-tRNA synthetase encoding gene was found in M. hominis. The G + C content of the gene was found to be 28.6%, which is significantly lower than in other prokaryotes. The gene was located 4 kb upstream of the M. hominis PG21 rRNA B operon.

Amino Acid Sequence↗
PubMed1994

The humoral immune response to Chlamydia trachomatis in patients with acute reactive arthritis.

Sera from 25 patients with clinical signs of reactive arthritis were analysed for antibodies against Chlamydia trachomatis by immunoblotting. Purified elementary bodies, purified Chlamydia outer membrane complexes, and purified recombinant subcomponents were used as antigens. Antibodies against C. trachomatis cysteine rich outer membrane protein 2 (Omp2) and lipopolysaccharide (LPS) were detected in 10 patients. Thus 40% of the patients presented antibodies specific for C. trachomatis. There was no correlation between acute reactive arthritis and antibodies to heat-shock proteins GroEL, GroES and DnaK.

Acute Disease↗
PubMed1994

Interaction of the Chlamydia trachomatis histone H1-like protein (Hc1) with DNA and RNA causes repression of transcription and translation in vitro.

The 18 kDa histone H1-like protein from Chlamydia trachomatis (Hc1) is a DNA-binding protein thought to be involved in condensation of the chlamydial chromosome during late stages in the chlamydial life cycle. Expression of Hc1 in Escherichia coli results in an overall relaxation of DNA and severely affects DNA, RNA and protein synthesis. We have analysed the interaction of Hc1 with single-stranded DNA and RNA by Southwestern and Northwestern blotting. Furthermore, we show that purified, recombinant Hc1 dramatically affects transcription and translation in vitro at physiologically relevant concentrations. These results were found to coincide with the formation of condensed Hc1-DNA and Hc1-RNA complexes as revealed by agarose gel electrophoresis and electron microscopy. The implications of these results for possible functions of Hc1 in vivo are discussed.

Chlamydia trachomatis↗
PubMed1994

Chlamydia trachomatis serovar L2 induces protein tyrosine phosphorylation during uptake by HeLa cells.

Chlamydia trachomatis L2 is an obligate intracellular microorganism with a unique biphasic life cycle. The extracellular form, the elementary body (EB), is infectious but metabolically inactive. Attachment of EBs to host cells is medicated by a heparan sulfate-like glycosaminoglycan. Following attachment, the EB is internalized within a membrane-bound vesicle, and during the first 8 h of infection the vesicles are transported to a perinuclear location where they aggregate and fuse. By use of a monoclonal antibody against phosphotyrosine, we showed that three classes of proteins are tyrosine phosphorylated: a triple band of 68, 66, and 64 kDa, a 97-kDa band, and a 140-kDa band. The phosphorylation could be detected by immunoblotting from 15 min after infection of HeLa cells. We followed the movement of the EBs and the tyrosine phosphorylation of proteins by double-labelling immunofluorescence microscopy with the same monoclonal anti-phosphotyrosine antibody and a polyclonal antibody against the C. trachomatis L2 outer membrane complex. During the first 8 h of infection, the phosphorylation colocalized with EBs. Sixteen hours after infection, EBs have reorganized to the replicating reticulate bodies, forming an inclusion. At this time, phosphorylation was seen as dotted spots in the periphery of the inclusion.

Actins↗
PubMed1994

Characterization of a linear epitope on Chlamydia trachomatis serovar L2 DnaK-like protein.

A cytoplasmic 75-kDa immunogen from Chlamydia trachomatis serovar L2 has previously been characterized as being similar to the Escherichia coli heat shock protein DnaK. We have localized a linear epitope for one monoclonal antibody specific for C. trachomatis DnaK. By use of a recombinant DNA technique, the epitope was limited to 14 amino acids. With synthetic peptides, the epitope was further limited to eight amino acids. Six of these amino acids are conserved in bovine HSP70, which has a known three-dimensional structure. The amino acid sequence homologous to the epitope is located in a linear part of the HSP70 molecule known as connect II.

Amino Acid Sequence↗
PubMed1994

Sequencing analysis reveals a unique gene organization in the gyrB region of Mycoplasma hominis.

The homolog of the gyrB gene, which has been reported to be present in the vicinity of the initiation site of replication in bacteria, was mapped on the Mycoplasma hominis genome, and the region was subsequently sequenced. Five open reading frames were identified flanking the gyrB gene, one of which showed similarity to that which encodes the LicA protein of Haemophilus influenzae. The organization of the genes in the region showed no resemblance to that in the corresponding regions of other bacteria sequenced so far. The gyrA gene was mapped 35 kb downstream from the gyrB gene.

Amino Acid Sequence↗
PubMed1994

Reciprocal exchange of minor components of type 1 and F1C fimbriae results in hybrid organelles with changed receptor specificities.

Type 1 and F1C fimbriae are surface organelles of Escherichia coli which mediate receptor-specific binding to different host surfaces. Such fimbriae are found on strains associated with urinary tract infections. The specific receptor binding of the fimbriae is due to the presence of receptor recognition proteins present in the organelles as minor structural elements. The organization of the fim and foc gene clusters encoding these fimbriae, as well as the structures of the organelles, are very similar, although the actual sequence homology of the structural elements is not remarkable; notably, the sequence identity between the minor components of the type 1 and F1C fimbriae is only 34 to 41%. Type 1 fimbriae mediate agglutination of guinea pig erythrocytes, whereas F1C fimbriae do not confer agglutination of any types of erythrocytes tested. However, F1C fimbriae mediate specific adhesion to epithelial cells in the collecting ducts of the human kidney as well as to cells of various cell lines. This report addresses the question of fimbrial promiscuity. Our data indicate that minor fimbrial structural elements can be exchanged between the two fimbrial systems, resulting in hybrid organelles with changed receptor specificity. This is the first study on reciprocal exchange of structural components from two different fimbrial systems.

Animals↗
PubMed1994

Detection of RET proto-oncogene point mutations in paraffin-embedded pheochromocytoma specimens by nonradioactive single-strand conformation polymorphism analysis and direct sequencing.

The suitability of formalin-fixed and paraffin-embedded tumor material was evaluated for molecular analysis of the RET proto-oncogene. We analyzed exons 10, 11, and 16 for point mutations in seven sporadic and six multiple endocrine neoplasia (MEN) 2A-associated pheochromocytomas by a nonradioactive single-strand conformation polymorphism assay followed by nonradioactive direct sequencing of PCR-amplified DNA using an automated DNA sequencer. All MEN 2A-associated pheochromocytomas contained a heterozygous missense germline mutation within cystine codons of the cysteine-rich extracellular domain encoded by exons 10 and 11. Mutations were located in codon 619 (TGC-->TCC; Cys-->Ser) in one, in codon 635 (TGC-->CGC; Cys--Arg) in three, and in codon 635 (TGC-->TAC; Cys-->Tyr) in two pheochromocytomas. No tumor-specific (somatic) mutations were detected in exons 10, 11, and 16 of the sporadic pheochromocytomas. These data support recent findings that germline point mutations that are clustered in distinct cysteine codons of the RET proto-oncogene are involved in the neoplastic phenotype of the MEN 2A syndrome. Our results demonstrate that both nonradioactive single-strand conformation polymorphism and direct sequencing are suitable methods to detect single base substitutions in DNA extracted from archival material.

Adolescent↗
PubMed1993

[Chlamydia pneumoniae].

Chlamydia are obligate intracellular bacteria with a biphasic life cycle. In 1986 a new species of Chlamydiae, Chlamydia psittaci TWAR was discovered, later named Chlamydia pneumoniae. C. pneumoniae is probably an important pathogen causing respiratory tract infections in man, ranging from subclinical infection to life threatening pneumonia. Infections can occur in epidemics. Furthermore, associations between C. pneumoniae infection and coronary artery disease, lethal myocarditis and sarcoidosis have been suggested. The current diagnostic method is based on detection of specific antibodies against C. pneumoniae in blood. Macrolides or tetracyclines are the drugs of choice for treatment of C. pneumoniae infection.

Chlamydia Infections↗
PubMed1993

Interaction between the Chlamydia trachomatis histone H1-like protein (Hc1) and DNA.

The gene encoding the Chlamydia trachomatis histone H1-like protein (Hc1) from serovar L2 was cloned into Escherichia coli by use of expression vector pET11d. In this vector, transcription of the gene is under the control of a bacteriophage T7 promoter, and T7 RNA polymerase is inducible in the host. Following induction, the E. coli cells were lysed gently. Gel filtration of the lysate revealed comigration of DNA and Hc1 in the voided volume. Electron microscopy revealed the DNA to be complexed with protein in large aggregates, often in the form of spherical bodies. Purified recombinant Hc1 maintained its DNA-binding capacity and was able at high concentrations to form condensed aggregates with DNA (one molecule of Hc1 per base pair) independently of the form or size of the DNA but with a slight preference for supercoiled DNA. Hc1 alone is thus able to package DNA into condensed spherical bodies.

Base Sequence↗
PubMed1992

Chlamydia trachomatis Mip-like protein.

A 27 kDa Chlamydia trachomatis Mip-like protein with homology of a 175-amino-acid C-terminal fragment to the surface-exposed Legionella pneumophila mip-gene product has previously been described. In this paper the entire chlamydia Mip-like sequence of C. trachomatis serovar L2 (lymphogranuloma venereum (LGV) biovar) is presented. The sequence shows high similarity to the legionella Mip protein and its C-terminal region, like that of the legionella Mip, has high amino acid similarity to eukaryotic and prokaryotic FK506-binding proteins. The chlamydial mip-like gene was detected by polymerase chain reaction (PCR) in other C. trachomatis serovars and by sequencing of the mip-like genes of serovars B and E (trachoma biovar) was shown to be highly conserved within the two major biovars of C. trachomatis. Monoclonal and polyclonal antibodies raised against the recombinant Mip-like protein failed to demonstrate surface-exposed epitopes on infectious elementary bodies or reproductive reticulate body forms either by immunofluorescence or immuno-gold electron microscopy. However, a complement-dependent inhibition of up to 91% of infectivity for cell cultures was observed with antibodies to the N-terminal fragment of the Mip-like protein suggesting that antibody-accessible epitopes are present on infectious EBs.

Amino Acid Sequence↗
PubMed1992

Physical and genetic mapping of the genomes of five Mycoplasma hominis strains by pulsed-field gel electrophoresis.

We present the complete maps of five Mycoplasma hominis genomes, including a detailed restriction map and the locations of a number of genetic loci. The restriction fragments were resolved by field inversion gel electrophoresis or by the contour-clamped homogeneous-electric-field system of pulsed-field gel electrophoresis. All the ApaI, SmaI, BamHI, XhoI, and SalI restriction sites (total of 21 to 33 sites in each strain) were placed on the physical map, yielding an average resolution of 26 kb. The maps were constructed using three different approaches: (i) size determination of DNA fragments partially or completely cleaved with one or two restriction enzymes, (ii) hybridization analysis with purified restriction fragments and specific probes, and (iii) use of linking clones. A genetic map was constructed by hybridization with gene-specific probes for rpoA, rpoC, rrn, tuf, gyrB, hup, ftsY, the unc operon, the genes for two M. hominis-specific antigenic membrane proteins, and one gene encoding a protein with some homology to Escherichia coli alanyl-tRNA synthetase. The positions of mapped loci were partially conserved in the five strains except in one strain in which a 300-kb fragment was inverted. The numbers and order of mapped restriction sites were only partly conserved, and this conservation was restricted to certain regions. The gene order was compared with the gene order established for other bacteria and was found to be identical to that of the phylogenetically related Clostridium perfringens. The genome size of the M. hominis strains varied from 704 to 825 kb.

Blotting, Southern↗
PubMed1991

Analysis of the nucleotide sequence of the Mycoplasma hominis tuf gene and its flanking region.

A gene from Mycoplasma hominis PG21 similar to the tuf gene encoding the elongation factor Tu (EF-Tu) of Escherichia coli was cloned and sequenced. The 1193-bp open reading frame flanked by a putative promoter and a potential stem-and-loop structure encoded a 44-kDa polypeptide. The tuf gene of M. hominis PG21 has the lowest G + C content seen in prokaryotes (38.2%). A gene (mhlmp1) encoding a variable surface exposed membrane protein (LMP1) was found downstream the 3' end of the tuf gene. It was found that the highly conserved tuf gene was linked to the highly variable mhlmp1 gene in 26 different M. hominis strains.

Antibodies, Monoclonal↗
PubMed1991

[The polymerase chain reaction used in microbiologic diagnosis].

Among the new DNA technological methods for microbiological diagnosis, the most promising seems to be the polymerase chain reaction (PCR). The sensitivity of the method is approximately 100% and the specificity is between 90% and 100%, depending on the availability of an already existing test sufficient for comparison. The PCR is able to detect only one microorganism in clinical samples. Because of this high sensitivity, contamination of samples is one of the major problems. After certain preparation procedures, the method can be used on different kinds of human sample material. The PCR could be used for detection of a microorganism, for detection of a single gene (toxin gene) or for typing of microorganisms. It is expected that, in the immediate future PCR will be a prominent part of the methods used in microbiological diagnosis.

Bacterial Infections↗
PubMed1991

Evaluation of urogenital Chlamydia trachomatis infections by cell culture and the polymerase chain reaction using a closed system.

Two hundred and fifty-four specimens from males and females consulting a clinic for sexually transmitted diseases were analyzed for genital Chlamydia trachomatis infection. Each clinical sample was tested by the cell culture technique and the polymerase chain reaction using a closed system. When the two test systems were compared, the overall sensitivity of the polymerase chain reaction was 96% and the specificity 94% when compared to the cell culture technique. By use of a closed system for DNA extraction and sample transfer for the polymerase chain reaction, contamination of the samples was minimized. The polymerase chain reaction detected a higher number of Chlamydia trachomatis infections among both symptomatic and asymptomatic females and males, and it also detected Chlamydia trachomatis at an earlier stage of infection when compared to cell culture. The polymerase chain reaction did not detect Chlamydia trachomatis after sufficient antibiotic treatment of the chlamydial infections.

Base Sequence↗
PubMed1991

Chlamydia trachomatis contains a protein similar to the Legionella pneumophila mip gene product.

A 27kDa Chlamydia trachomatis L2 protein was characterized by the use of monoclonal antibodies and by two-dimensional gel electrophoresis. The protein was shown to be located in the membrane of reticulate bodies as well as elementary bodies. Its synthesis could be detected from 10 hours post-infection. Cloning and sequence analysis of the distal part of the gene revealed an open reading frame of 175 amino acids. Comparison of the deduced amino acid sequence with the NBRF data base revealed significant homology between the 27 kDa chlamydial membrane protein and the product of the macrophage infectivity potentiator (mip) gene of Legionella pneumophila.

Amino Acid Sequence↗