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Biomedical subjects

J London

Publications and source records attributed to J London.

At least 55 records · Page 3Linked to original sources
PubMed1990

Structure of a streptococcal adhesin carbohydrate receptor.

Interactions between complementary protein and carbohydrate structures on different genera of human oral bacteria have been implicated in the formation of dental plaque. The carbohydrate receptor on Streptococcus sanguis H1 (one of the primary colonizing species) that is specific for the adhesin on Capnocytophaga ochracea ATCC 33596 (a secondary colonizer) has been isolated from the streptococcal cell wall, purified, and structurally characterized. The hexasaccharide repeating unit of the polysaccharide was purified by reverse-phase, amino-bonded silica, and gel permeation high performance liquid chromatography. Earlier studies established that the repeating unit was a hexasaccharide composed of rhamnose, galactose, and glucose in the ration of 2:3:1, respectively. In the present study, determination of absolute configuration by gas chromatography of the trimethylsilyl (+)-2-butyl glycosides revealed that the rhamnose residues were of the L configuration while the hexoses were all D. 252Californium plasma desorption mass spectrometry of the native, the acetylated and the reduced and acetylated hexasaccharide determined that the molecular mass of the native hexasaccharide was 959, and that the 2 rhamnose residues were linked to each other at the nonreducing terminus of the linear molecule. Methylation analysis revealed the positions of the glycosidic linkages in the hexasaccharide and showed that a galactose residue was present at the reducing end. The structural characterization of the hexasaccharide was completed by one and two dimensional 1H and 13C NMR spectroscopy. Complete 1H and 13C assignments for each glycosyl residue were established by two-dimensional (1H,1H) correlation spectroscopy, homonuclear Hartmann-Hahn, and (13C,1H) correlation experiments. The configurations of the glycosidic linkages were inferred from the chemical shifts and coupling constants of the anomeric 1H and 13C resonances. The sequence of the glycosyl residues was determined by a heteronuclear multiple bond correlation experiment. These data show that the structure of the hexasaccharide repeating unit derived from the cell wall polysaccharide of S. sanguis H1 is: alpha-L-Rhap-(1----2)-alpha-L-Rhap-(1----3)-alpha-D-Galp- (1----3)-beta-D-Galp-(1----4)-beta-D-Glcp-(1----3)-alpha/beta-D-Gal.

Bacterial Adhesion↗
PubMed1990

A novel gene member of the human glycophorin A and B gene family. Molecular cloning and expression.

A new gene closely related to the glycophorin A (GPA) and glycophorin B (GPB) genes has been identified in the normal human genome as well as in that of persons with known alterations of GPA and/or GPB expression. This gene, called glycophorin E (GPE), is transcribed into a 0.6-kb message which encodes a 78-amino-acid protein with a putative leader peptide of 19 residues. The first 26 amino acids of the mature protein are identical to those of M-type glycophorin A (GPA), but the C-terminal domain (residues 27-59) differs significantly from those of glycophorins A and B (GPA and GPB). The GPE gene consists of four exons distributed over 30 kb of DNA, and its nucleotide sequence is homologous to those of the GPA and GPB genes in the 5' region, up to exon 3. Because of branch and splice site mutations, the GPE gene contains a large intron sequence partially used as exons in GPA and GPB genes. Compared to its counterpart in the GPB gene, exon 3 of the GPE gene contains several point mutations, an insertion of 24 bp, and a stop codon which shortens the reading frame. Downstream from exon 3, the GPE and the GPB sequences are virtually identical and include the same Alu repeats. Thus, it is likely that the GPE and GPB genes have evolved by a similar mechanism. From the analysis of the GPA, GPB and GPE genes in glycophorin variants [En(a-), S-s-U- and Mk], it is proposed that the three genes are organized in tandem on chromosome 4. Deletion events within this region may remove one or two structural gene(s) and may generate new hybrid structures in which the promoter region of one gene is positioned upstream from the body of another gene of the same family. This model of gene organization provides a basis with which to explain the diversity of the glycophorin gene family.

Amino Acid Sequence↗
PubMed1990

Uncommon pathways of metabolism among lactic acid bacteria.

A small number of lactic acid bacteria possess the ability to derive energy from organic molecules not utilized by the vast majority of representatives of this large group of microorganisms. Thus, strains of Lactobacillus casei and enterococci readily grow at the expense of substrates such as gluconate, malate and pentitols. Transport of gluconate and pentitols is catalysed by phosphotransferase systems unique to these bacteria. Similarly, the initial steps in pentitol dissimilation are mediated by enzymes found only in Lb. casei and Streptococcus avium.

Amino Acids↗
PubMed1990

Purification and characterization of a Bacteroides loeschei adhesin that interacts with procaryotic and eucaryotic cells.

The adhesin of Bacteroides loeschei PK1295 that mediates coaggregation with Streptococcus sanguis 34 and hemagglutination of erythrocytes was purified to electrophoretic homogeneity. The lectinlike protein has an estimated native Mr of 450,000 and consists of six subunits of identical molecular weight (Mr 75,000). The purified adhesin appears to be a basic protein with a pI between 7.4 and 8.0. Amino acid and N-terminal sequence analyses were carried out with the purified protein. These indicated that the protein contains a large number of Asx and Glx residues as well as basic amino acid residues. The binding site of the pure adhesin retained its native configuration during purification. When preincubated with streptococcal partner cells at pH 4.6, the adhesin prevented B. loeschei cells from coaggregating with the streptococci. An adhesin preparation adjusted to a pH of 6.8 rapidly agglutinated both streptococci and neuraminidase-treated erythrocytes. Galactosides inhibited the agglutination reactions.

Adhesins, Bacterial↗
PubMed1990

Buspirone in depressed outpatients: a controlled study.

One hundred fifty-five outpatients suffering from major depression with significant anxiety entered a double-blind study comparing 8 weeks of treatment with buspirone or placebo. Twenty-nine percent of buspirone and 40 percent of placebo patients discontinued treatment before 8 weeks. Major efficacy measures were the Hamilton Rating Scale for Depression (HAM-D) total score, the HAM-D retardation and anxiety factors, the HAM-D Rickels and Bech core depression clusters, the Clinical Global Impressions (CGI), and the Hopkins Symptom Checklist (HSCL). Results were consistent across all outcome measures, including the two core depression clusters, with treatment response to buspirone significantly better than to placebo at treatment endpoint. Seventy percent of buspirone and 35 percent of placebo patients (p less than .01) were rated moderately or markedly improved after 8 weeks of therapy. Buspirone was found to be safe and well-tolerated by patients with major depression and concomitant anxiety at doses of up to 90 mg/day.

Anxiety Disorders↗
PubMed1990

A placebo-controlled study of enciprazine in the treatment of generalized anxiety disorder: a preliminary report.

Enciprazine is a propanolamine derivative with a preclinical profile similar to buspirone but with less affinity for the postsynaptic dopamine receptor (Linden et al. 1988). We report on the outcome, using intent-to-treat data, of a 5-week, double-blind trial comparing three dose strengths of enciprazine (5 mg t.i.d., 10 mg t.i.d., and 20 mg t.i.d.) to placebo. A dose escalation was permitted after 2 weeks of active drug treatment, which 61 percent of patients overall took advantage of. A "last observation carried forward" (LOCF) analysis found a mean improvement in Hamilton Anxiety Scale (HAM-A) scores by Week 5 of -11.0 for the combined enciprazine treatment groups, and -4.4 for the placebo group (p less than .05). Fifty-two percent of enciprazine patients were judged to be "much" or "very much" improved, whereas none of the placebo patients were judged to have comparable improvement. Enciprazine was well-tolerated, with low levels of sedative and asthenic side effects reported. The compound appears to have promise as an anxiolytic agent.

Adult↗
PubMed1989

Structure of the 5' flanking region of the gene encoding human glycophorin A and analysis of its multiple transcripts.

Glycophorin A (GPA), the major sialoglycoprotein of human erythrocytes, is the carrier for blood group MN antigens and a receptor for viruses, bacteria and parasites. (1) Three distinct GPA mRNAs (1.0, 1.7 and 2.2 kb) have been previously identified in erythroid tissues by Northern-blot analysis. It is shown here by sequence analysis of several human fetal liver cDNAs, and by transcription start point (tsp) determination using primer extension analysis, that the production of the multiple GPA mRNAs is governed by poly(A) site choice generating 3'-untranslated regions of different length, and not by the tsp heterogeneity, since all messages exhibit the same cap site (tsp). (2) The structural gene encoding GPA has been recently cloned [Vignal et al., Eur. J. Biochem. 184 (1989) 337-344; Kudo and Fukuda, Proc. Natl. Acad. Sci. USA 86 (1989) 4619-4623] and we have now determined the sequence of a DNA genomic fragment upstream from the tsp. This fragment does not contain the typical TATA and CAAT boxes found in a number of tissue-specific genes, but contains typical motifs like the CACC, nuclear factor erythroid 1 and 2 elements, which have been identified recently in several erythroid-specific promoters, therefore suggesting that transcription of these genes might be regulated by the same or analogous factors.

Amino Acid Sequence↗
PubMed1989

Molecular analysis of glycophorin A and B gene structure and expression in homozygous Miltenberger class V (Mi. V) human erythrocytes.

In the Miltenberger class V (Mi. V) condition, red cells lack glycophorin A (GPA) and glycophorin B (GPB) but carry instead an unusual glycoprotein thought to be a hybrid molecule produced by the unequal crossing-over between the closely linked genes encoding for GPA and GPB. By Western blot analysis with rabbit anti-GPA antibodies specific for discrete domains of GPA, it was found that the Mi. V glycoprotein (donor F. M.) contains approximately 60 amino acid residues of GPA at its N-terminus. As a preliminary approach to the molecular analysis of this variant the restriction maps of the GPA and GPB genes were established by Southern blot analysis of genomic DNA and from genomic clones isolated from a human leukocyte library constructed in lambda EMBL4. The GPA and GPB genes cover about 30 kb of DNA and are organized into seven exons (A-1-A-7) and five exons (B-1-B-5), respectively. In addition to the normal genes, a third gene (named inv), closely resembling the GPA and GPB genes, was also identified. In the homozygous Mi. V individual the normal GPA and GPB genes were absent, but an unusual form of gene structure was detected by Southern blot analysis. The Mi. V glycoprotein gene was composed of exon B-1 of the GPB gene followed by exons A-2 and A-3 of the GPA gene and the exons B-3, B-4 and B-5 of the GPB gene. Exon B-1 can be distinguished from exon A-1 of GPA since it is located within a different restriction fragment, but both encode the same amino acid sequence (N-terminal region of the signal peptides). Using the polymerase chain reaction, the junction between exon A-3 and exon B-3 was confirmed by amplification of the DNA region where the putative crossing-over has occurred and it was deduced that the Mi. V glycoprotein is a hybrid molecule composed of amino acid residues 1-58 from GPA fused to amino acid residues 27-72 of GPB. In addition, the finding that part of the signal peptide and the 5'-untranslated region are derived from GPB suggests that the genetic background of the Mi. V variant is rather complex and may involve a cascade of recombination or gene conversion events.

Amino Acid Sequence↗
PubMed1989

Human erythrocyte glycophorin C. Gene structure and rearrangement in genetic variants.

We have previously shown that a deletion of approximately 3 kilobases in the unique glycophorin C (GPC) gene, which encodes for the human erythrocyte glycophorins C and D, is associated with the Gerbich (Ge) blood group deficiency (Ge-2,-3 and Ge-2,+3 types) (Le van Kim, C., Colin, Y., Blanchard, D., Dahr, W., London, J. & Cartron, J.P. (1987) Eur. J. Biochem. 165, 571-579). We have now isolated and characterized the structure of the GPC gene from the common Ge+2,+3 donors and from a Ge-2,-3 variant (Ge-2,-3 gene). The GPC gene is organized in four exons distributed over 13.5-kilobase pairs (kbp) DNA and contains two directly repeated domains of 3.4 kbp in length which are likely derived from the recent duplication of a unique ancestral domain. Restriction mapping and sequence analysis indicate that a 3.4-kbp deletion within this gene, arising probably by unequal crossing over between the two repeated domains, is responsible for the formation of the Ge-2,-3 gene. The breakpoints of the deletion are located within introns 2 and 3, and therefore exon 3 is removed. The defective gene is transcribed as a mRNA with a continuous open reading frame extending over 300 nucleotides which is translated into an unusual sialoglycoprotein present on Ge-2,-3 red cells. The primary structure of this new glycoprotein has been deduced from nucleotide sequencing. It is proposed in addition, that another 3.4-kb deletion within the GPC gene eliminates exon 2 only by a similar mechanism and generates a defective gene encoding for the abnormal glycoprotein present on Ge-2,+3 erythrocytes. Interestingly, the same deletion which lead to the rare Ge-2,-3 genetic condition, occurred spontaneously and frequently in the cloned GPC gene during the propagation of the recombinant phages in Escherichia coli. From these observations we suggest that the Ge-2,-3 and Ge-2,+3 genes might represent the two allelic forms of a unique ancestral form of the GPC gene, following successive internal duplication and deletion events.

Amino Acid Sequence↗
PubMed1989

Use of adhesin-specific monoclonal antibodies to identify and localize an adhesin on the surface of Capnocytophaga gingivalis DR2001.

Monoclonal antibodies capable of inhibiting coaggregation between Capnocytophaga gingivalis DR2001 and Actinomyces israelii PK16 were used to identify the adhesin on C. gingivalis that mediates the interaction. The monoclonal antibodies were used to demonstrate that a 140-kilodalton polypeptide found in the outer membrane of C. gingivalis was the adhesin responsible for coaggregation. A coaggregation-defective mutant that was unable to coaggregate with A. israelii lacked this large polypeptide. The monoclonal antibodies were also used to estimate the number of binding sites on the surfaces of individual cells and show how the adhesin molecules were arranged on the outer membrane. Values of between 220 and 280 were obtained for the number of adhesin molecules per cell. Immunoelectron microscopy performed with the monoclonal antibodies revealed that the adhesin molecules were arranged nonuniformly on the bacterial surface and occurred singly, in pairs, and in small clusters.

Actinomyces↗
PubMed1989

Bacteroides loeschei PK1295 cells express two distinct adhesins simultaneously.

Bacteroides loeschei synthesizes two distinct adhesins that mediate its coaggregation with Streptococcus sanguis 34 and Actinomyces israeli PK14. Streptococcal adhesin-specific and actinomyces adhesin-specific monoclonal antibodies were used to prepare antibody-coated 5- or 10-nm gold particles. These were used in immunoelectron microscopic studies to establish that essentially all bacteroides cells in a population express both adhesins. In general, the two sizes of gold particles representing each type of adhesin appeared to be spatially separated on neighboring fimbriae of B. loeschei. Deposition of antibody-coated gold particles, representing both types of adhesin, at or near the same fimbria was observed less frequently.

Antibodies, Monoclonal↗
PubMed1989

Fimbria-associated adhesin of Bacteroides loeschei that recognizes receptors on procaryotic and eucaryotic cells.

Inhibition studies with a set of adhesin-specific monoclonal antibodies and various sugars revealed that a fimbria-associated adhesin of Bacteroides loeschei recognizes receptors on both procaryotic and eucaryotic cells. These interactions permit this bacterium to attach to both types of cells, producing coaggregates in the presence of strains of Streptococcus sanguis and hemagglutination in the presence of neuraminidase-treated human erythrocytes.

Actinomyces↗
PubMed1989

Isolation of a coaggregation-inhibiting cell wall polysaccharide from Streptococcus sanguis H1.

Coaggregation between Streptococcus sanguis H1 and Capnocytophaga ochracea ATCC 33596 cells is mediated by a carbohydrate receptor on the former and an adhesin on the latter. Two methods were used to release the carbohydrate receptor from the gram-positive streptococcus, autoclaving and mutanolysin treatment. The polysaccharide released from the streptococcal cell wall by either treatment was purified by ion-exchange chromatography; this polysaccharide inhibited coaggregation when preincubated with the gram-negative capnocytophaga partner. After hydrolysis of the polysaccharide by hydrofluoric acid (HF), the major oligosaccharide of the polysaccharide was purified by high-performance liquid chromatography. By analysis of the HF hydrolysis of the polysaccharide and the purified oligosaccharide, this major oligosaccharide appeared to be the repeating unit of the polysaccharide, with minor components resulting from internal hydrolysis of the major oligosaccharide. Gas chromatography results showed that the oligomer was a hexasaccharide, consisting of rhamnose, galactose, and glucose, in the ratio of 2:3:1, respectively. By weight, the purified hexasaccharide was a fourfold-more-potent inhibitor of coaggregation than the native polysaccharide. Resistance to hydrolysis by sulfuric acid alone and susceptibility to hydrolysis by HF suggested that oligosaccharide chains of the polysaccharide are linked by phosphodiester bonds. Studies with a coaggregation-defective mutant of S. sanguis H1 revealed that the cell walls of the mutant contained neither the polysaccharide nor the hexasaccharide repeating unit. The purification of both a polysaccharide and its constituent hexasaccharide repeating unit, which both inhibited coaggregation, and the absence of this polysaccharide or hexasaccharide on a coaggregation-defective mutant strongly suggest that the hexasaccharide derived from the polysaccharide functions as the receptor for the adhesin from C. ochracea ATCC 33596.

Adhesins, Bacterial↗
PubMed1989

Optical measurement of action potential activity in invertebrate ganglia.

Optical monitoring methods have reached the level of development where activity from a network of cells can be recorded in a minimally-dissected behaving animal. The spike activity in the buccal ganglion of Navanax was monitored during feeding and activity in the Aplysia abdominal ganglion was monitored during the gill-withdrawal reflex. Approximately 30 neurons in the Navanax buccal ganglion were active during feeding and between 250 and 400 neurons in the Aplysia abdominal ganglion were active during the gill-withdrawal reflex. A reasonably complete understanding of the neuronal basis of the gill withdrawal may not be possible with presently available scientific methods. Substantial improvements in signal-to-noise ratio in optical measurements will be necessary before the majority of synaptic potentials can be detected optically. Understanding circuits that involve more than a few neurons will be a challenge to neurobiologists.

Action Potentials↗
PubMed1988

Alteration of the genes for glycophorin A and B in glycophorin-A-deficient individuals.

Glycophorins A and B are homologous glycoproteins of the red cell membrane which carry the blood-group MN and Ss antigens, respectively, and are encoded by two distinct genes closely linked on chromosome 4, which are probably derived from each other by duplication during evolution. The lack of glycophorin A is associated with the rare phenotype En(a-), indicating individuals who are defective for MN antigens, as well as for the Ena antigens, also located on this glycoprotein. The En(a-) condition is heterogenous and includes two categories of variants exemplified by the Finnish and the English types referred to as En(Fin) and En(UK), respectively. By Southern blot and preliminary genomic clone analyzes we have compared the status of the genes for glycophorins A and B, as well as that of the gene encoding glycophorin C, another unrelated red cell membrane glycoprotein, in the En(a-) variants and in the En(a+) control donors. Our data indicate that the En(Fin) variant is homozygous for a complete deletion of the glycophorin A gene without any detectable abnormality of the genes encoding glycophorins B or C. In the genome of the En(UK) variant, with the presumed genotype Mk/En(UK), and where the Mk condition abolishes the expression of MN and Ss antigens, we have identified several abnormalities of the glycophorin A and B genes, but the glycophorin C gene was unaffected. Our results strongly support the view that in Mk chromosome the glycophorin A and B genes are largely deleted, whereas the En(UK) chromosome probably contains a gene fusion product encoding a hybrid glycoprotein AM-B, composed of the N-terminal portion of a blood group M-type glycophorin A and of the C-terminal portion of glycophorin B. The determination of the 5' and 3' limits of the hybrid gene and elucidation of the mechanism involved will require sequencing of the rearranged DNA of the variant and a full knowledge of the organization of the glycophorin A and B genes.

Blotting, Southern↗
PubMed1988

Characterization of cDNA clones for human glycophorin A. Use for gene localization and for analysis of normal of glycophorin-A-deficient (Finnish type) genomic DNA.

Glycophorin A is the major membrane sialoglycoprotein of human erythrocytes and represents a typical example of a transmembrane glycoprotein. The functional role of this cell-surface component is not known but it represents a receptor for viruses, bacteria and parasites like Plasmodium falciparum. 1. Two cDNA clones encoding glycophorin A have been characterized from human fetal cDNA libraries. The longer cDNA extended from the coding region of glycophorin A (residues 4-131) to the 3' untranslated region which included two polyadenylation signals and a poly(A) tail. 2. The structural gene for glycophorin A is located on chromosome 4, q28-q31 as shown by in situ hybridization, thus confirming the previous localization by genetic linkage analysis. 3. Three distinct mRNA species (1.0 kb, 1.7 kb and 2.2 kb) have been identified in erythroid spleen. Northern blot analyses with a probe directed against the 3' untranslated region of the mRNAs indicated that all these species share a homologous 3' non-coding region and that the first polyadenylation signal downstream the stop codon is not used. 4. Preliminary studies by Southern blot analysis of the genomic DNA from normal En(a+) and rare En(a-) donors suggest that the glycophorin A gene has a complex organization and is largely deleted in donors of the En(a-) phenotype (Finnish type) who lack glycophorin A on their red cells.

Base Sequence↗