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S Scherer

Publications and source records attributed to S Scherer.

At least 91 records · Page 5Linked to original sources
PubMed1996

A single center evaluation of the Collaborative Transplant Study (CTS) DNA project.

Historical HLA class II serological typing results of transplantations performed in "The Leuven Collaborative Group for Transplantation" were subjected to retrospective Restriction Fragment Length Polymorfism (RFLP) DNA control typing by the Collaborative Transplant Study (CTS) DNA project using Polymerase Chain Reaction (PCR)-based DNA methods. We re-evaluated the serology/ RFLP-discrepant CTS DNA data for our local patients transplanted during a historical period (January 1988 until May 1992) before any class II DNA typing was performed in our tissue typing laboratory. These retyping results confirm both the CTS data for patient typing and the Eurotransplant data for donor typing. A confirmed high discrepancy rate of 19.0% (after exclusion of 2.2% transcription errors) was found in the patient population. A low discrepancy rate of 6.8% (after exclusion of 2.2% transcription errors) for the donor population is concordant with the Eurotransplant donor data. Only 4 of the 588 individuals were found to be incorrectly typed by the RFLP method; all involving the specificities DRB1*1102. This indicates that RFLP typing, as performed by the CTS DNA project, can be considered a valid, retrospective DNA typing system for the accurate interpretation of class II matching in organ transplantation. A second conclusion to be drawn from this study is the need for prospective DNA typing for kidney transplant recipients, as the discrepancy rate in this cohort is high. Our results suggest that with good quality serological HLA-DR typing, prospective donor DNA typing is not urgently needed.

Belgium↗
PubMed1996

Construction of luciferase reporter bacteriophage A511::luxAB for rapid and sensitive detection of viable Listeria cells.

Specific transfer and expression of bacterial luciferase genes via bacteriophages provides an efficient way to detect and assay viable host cells. Listeria bacteriophage A511 is a genus-specific, virulent myovirus which infects 95% of Listeria monocytogenes serovar 1/2 and 4 cells. We constructed recombinant derivative A511::luxAB, which carries the gene for a fused Vibrio harveyi LuxAB protein inserted immediately downstream of the major capsid protein gene (cps). Efficient transcription is initiated by the powerful cps promoter at 15 to 20 min postinfection. Site-specific introduction of the luciferase gene into the phage genome was achieved by homologous recombination in infected cells between a plasmid carrying A511 DNA flanking luxAB and phage DNA. Recombinants occurred in the lysate at a frequency of 5 x 10(-4) and were readily identified by the bioluminescent phenotype conferred on newly infected host cells. A511::luxAB can be used to directly detect Listeria cells. Following infection and a 2-h incubation period, numbers as low as 5 x 10(2) to 10(3) cells per ml were detected by using a single-tube luminometer. Extreme sensitivity was achieved by including an enrichment step prior to the lux phage assay; under these conditions less than 1 cell of L. monocytogenes Scott A per g of artificially contaminated salad was clearly identified. The assay is simple, rapid, inexpensive, and easy to perform. Our findings indicate that A511::luxAB is useful for routine screening of foods and environmental samples for Listeria cells.

Bacteriophages↗
PubMed1996

Nucleotide sequence and taxonomical distribution of the bacteriocin gene lin cloned from Brevibacterium linens M18.

Linocin M18 is an antilisterial bacteriocin produced by the red smear cheese bacterium Brevibacterium linens M18. Oligonucleotide probes based on the N-terminal amino acid sequence were used to locate its single copy gene, lin, on the chromosomal DNA. The amino acid composition, N-terminal sequence, and molecular mass derived from the nucleotide sequence of an open reading frame of 798 nucleotides coding for 266 amino acids found on a 3-kb BamHI restriction fragment correspond closely to those obtained from the purified protein (N. Valdés-Stauber and S. Scherer, Appl. Environ. Microbiol. 60:3809-3814, 1994). No sequence homology to any protein or nucleotide sequences deposited in databases was found. Comparison of the nucleotide sequence and the N-terminal amino acid sequence derived from the protein suggests that B. linens M18 produces an N-formyl-methionyl-CAC tRNA. A wide taxonomical distribution of the gene within coryneform bacteria has been demonstrated by PCR amplification. The structural gene from linocin M18 is present at least in three Brevibacterium species, five Arthrobacter species, and five Corynebacterium species.

Actinomycetales↗
PubMed1996

Modified Listeria bacteriophage lysin genes (ply) allow efficient overexpression and one-step purification of biochemically active fusion proteins.

Listeria bacteriophage lytic enzymes are useful for in vitro applications such as rapid, gentle cell disruption, and they provide new approaches as selective antimicrobial agents for destruction of Listeria monocytogenes in contaminated foods. We describe here the amino-terminal modification of three cloned Listeria phage lysin genes (ply), resulting in fusion proteins with a 12-amino-acid leader containing six consecutive histidine residues. The recombinant enzymes retain their native specific activity and can be efficiently overproduced in Escherichia coli. By one-step metal chelate affinity chromatography, active lysins could be purified to more than 90% homogeneity.

Amino Acid Sequence↗
PubMed1996

Identification and purification of a family of dimeric major cold shock protein homologs from the psychrotrophic Bacillus cereus WSBC 10201.

Whole-cell protein patterns of a psychrotrophic Bacillus cereus strain from cultures grown at 7 and 30 degrees C were compared. This analysis revealed that at least three major proteins are expressed at a significantly higher rate at 7 degrees C than at 30 degrees C. The most abundant of these cold-induced proteins was a small polypeptide of 7.5 kDa, designated CspA, of B. cereus. In addition, four small proteins very similar in size to CspA were seen on both 7 degrees C and 30 degrees C two-dimensional protein gels. Immunoblot analysis using B. cereus anti-CspA antibodies indicated that the five proteins described above plus an additional sixth protein not visible on silver-stained two-dimensional gels are members of a B. cereus cold shock protein family. This hypothesis was corroborated by cloning and sequencing of the genes encoding five proteins of this family. The protein sequences deduced are highly similar and show homology to small procaryotic cold shock proteins and to the cold shock domain of eucaryotic Y-box proteins. Besides CspA, only one of the additional five CspA homologs was slightly cold inducible. In the presence of 100 mM NaCl, the two purified members of the protein family (CspA and CspE) elute as dimers at an apparent molecular mass of 15 kDa from a gel filtration column. At higher salt concentrations, they dissociate into their monomers. Their ability to bind to the ATTGG motif of single-stranded oligonucleotides was demonstrated by band shift analysis.

Adaptation, Physiological↗
PubMed1995

Structure of a novel oligosaccharide-mycosporine-amino acid ultraviolet A/B sunscreen pigment from the terrestrial cyanobacterium Nostoc commune.

Water-soluble UV-A/B-absorbing pigments are secreted by cells of the cosmopolitan terrestrial cyanobacterium Nostoc commune. The pigments constitute a complex mixture of monomers with molecular masses of up to 1801 Da. Two different chromophores with absorption maxima at 312 and 335 nm are linked to different amino acids and to oligosaccharides consisting of galactose, glucose, xylose, glucuronic acid, and glucosamine. The 335 nm chromophore is a 1,3-diaminocyclohexen derivative, while the chromophore with an absorption maximum at 312 nm is most likely a 3-aminocyclohexen-1-one derivative. These UV-inducible substances are the first mycosporines to be described covalently linked to oligosaccharides. The pigments are located in the extracellular glycan sheath of Nostoc colonies, where they form complexes of extremely high molecular mass that are attached noncovalently to the glycan sheath. Pigments occur in concentrations that permit the cells to attenuate a significant part of incident UV-B radiation.

Amino Acids↗
PubMed1995

Expression and regulation of kainate and AMPA receptors in uncommitted and committed neural progenitors.

Here we review experimental evidence of non-NMDA glutamate receptor expression in the embryonic central nervous system. AMPA- and kainate-preferring glutamate receptor subunit mRNA transcripts are detected in embryonic neurons, glia and neural progenitors. Functional assays demonstrate that in some cell subpopulations ionotropic glutamate receptors are expressed by progenitors before synapse formation and terminal differentiation, and may be present before lineage determination is specified. The activation of these receptors triggers induction of immediate early gene transcription in progenitor cells. The cloning and transcriptional analysis of upstream regulatory regions of glutamate receptor genes governing their temporal and tissue-specific expression are also discussed.

Animals↗
PubMed1995

Characterization of cryptic prophages (monocins) in Listeria and sequence analysis of a holin/endolysin gene.

Monocins in Listeria were induced by UV-irradiation of liquid cultures, and defective phage particles were purified from the lysates. Electron microscopy showed flexible, non-contractile bacteriophage-tail-like particles, consisting of specific proteins of molecular mass 20-45 kDa and pI 4.6-6.7. These particles were able to lyse listerial cells. DNA sequence homologies between chromosomal DNA of monocin-producing strains and labelled Listeria phage DNAs were inferred from DNA/DNA hybridizations, suggesting that most of the prophage DNA is still present in the listerial chromosome. An endolysin gene cpl2438 was cloned from listerial chromosomal DNA and was identified by its expression of lytic activity against Listeria cells in a bioassay. The gene consists of 864 nt encoding a protein of 287 aa with a calculated molecular mass of 32975 Da (CPL2438). This is in good agreement with the size of a protein observed in SDS-PAGE after overexpression of the lytic protein in Escherichia coli. The nucleotide sequence of a putative holin gene (hol2438, 291 nt) upstream of cpl2438 was determined after PCR-amplification of listerial DNA and it shows typical features common to the holin gene family. Expression of the encoded protein (HOL2438, 95 aa, 10.1 kDa) in E. coli was found to be lethal for the host cells. The results underline the close relationship between monocins and intact Listeria bacteriophages, indicating that monocins are incompletely assembled phage particles derived from cryptic prophages of Listeria, probably including the phage lysin.

Amino Acid Sequence↗
PubMed1995

A solitary plasmacytoma of donor origin arising 14 years after kidney allotransplantation.

Allotransplantation of solid organs transfers passenger leucocytes which may give rise to a state of persistent microchimaerism. In this report we describe the case of a patient who developed a solitary plasmacytoma in a transplanted kidney more than 10 years after allografting. The diagnosis was established on the basis of the presence of a monoclonal IgG kappa peak in the serum, and light chain proteinuria, the plasmacytoid features of tumour cells including cell surface expression of IgG, kappa light chains, CD20, CD38 and CD56, the absence of lytic bone lesions and a normal bone marrow biopsy, and the disappearance of the monoclonal IgG peak after graft nephrectomy. A donor origin of the tumour was established by HLA DNA typing of tumour, tumour-free kidney tissue, and peripheral blood leucocytes, respectively.

Adult↗
PubMed1995

Candida albicans ALS1: domains related to a Saccharomyces cerevisiae sexual agglutinin separated by a repeating motif.

Transfer of budding Candida albicans yeast cells from the rich, complex medium YEPD to the defined tissue culture medium RPMI 1640 (RPMI) at 37 degrees C and 5% CO2 causes rapid onset of hyphal induction. Among the genes induced under these conditions are hyphal-specific genes as well as genes expressed in response to changes in temperature, CO2 and specific media components. A cDNA library constructed from cells incubated for 20 min in RPMI was differentially screened with yeast (YEPD)- and hyphal (RPMI)-specific probes resulting in identification of a gene expressed in response to culture conditions but not regulated by the yeast-hyphal transition. The deduced gene product displays significant identity to Saccharomyces cerevisiae alpha-agglutinin, encoded by AG alpha 1, an adhesion glycoprotein that mediates mating of haploid cells. The presence of this gene in C. albicans is curious since the organism has not been observed to undergo meiosis. We designate the C. albicans gene ALS1 (for agglutinin-like sequence). While the N- and C-termini of the predicted 1260-amino-acid ALS1 protein resemble those of the 650-amino-acid AG alpha 1, ALS1 contains a central domain of tandem repeats consisting of a highly conserved 36-amino-acid sequence not present in AG alpha 1. These repeats are also present on the nucleotide level as a highly conserved 108 bp motif. Southern and Northern blot analyses indicate a family of C. albicans genes that contain the tandem repeat motif; at least one gene in addition to ALS1 is expressed under conditions similar to those for ALS1 expression. Genomic Southern blots from several C. albicans isolates indicate that the number of copies of the tandem repeat element in ALS1 differs across strains and, in some cases, between ALS1 alleles in the same strain, suggesting a strain-dependent variability in ALS1 protein size. Potential roles for the ALS1 protein are discussed.

Agglutinins↗
PubMed1995

Heterogeneous endolysins in Listeria monocytogenes bacteriophages: a new class of enzymes and evidence for conserved holin genes within the siphoviral lysis cassettes.

Listeria monocytogenes bacteriophages A118, A500 and A511 are members of three distinct phage groups with characteristic host ranges. Their endolysin (ply) genes were cloned and expressed in Escherichia coli as demonstrated by the conferred lytic phenotype when colonies of recombinant cells were overlaid with a lawn of Listeria cells. The nucleotide sequences of the cloned DNA fragments were determined and the individual enzymes (PLY118, 30.8 kDa; PLY500, 33.4 kDa; PLY511, 36.5 kDa) were shown to have varying degrees of homology within their N-terminal or C-terminal domains. Transcriptional analysis revealed them to be 'late' genes with transcription beginning 15-20 min post-infection. The enzymes were overexpressed and partially purified and their individual specificities examined. When applied exogenously, the lysins induced rapid lysis of Listeria strains from all species but generally did not affect other bacteria. Using hydrolysis of purified listerial cell walls, PLY511 was characterized as an N-acetylmuramoyl-L-alanine amidase (EC 3.5.1.28) and shows homology in its N-terminal domain to other enzymes of this type. In contrast, PLY118 and PLY500 were shown to represent a new class of cell wall lytic enzymes which cleave between the L-alanine and D-glutamate residues of listerial peptidoglycan; these were designated as L-alanoyl-D-glutamate peptidases. These two enzymes share homology in the N-terminal domain which we propose determines hydrolytic specificity. Highly conserved holin (hol) gene sequences are present upstream of ply118 and ply500. They encode proteins of structural similarity to the product of phage lambda gene S, and are predicted to be membrane proteins which form pores to allow access of the lysins to their peptidoglycan substrates. This arrangement of conserved holin genes with downstream lysin genes among the siphoviral lysis cassettes explains why the cytoplasmic endolysins alone are not lethal, since they require a specific transport function across the cell membrane.

Amino Acid Sequence↗