Effect of pregnancy-specific β1-glycoprotein on indoleamine-2,3-dioxygenase activity in human monocytes

The role of heterogenic human pregnancy-specific glycoprotein (PSG), obtained by the authors’ technology, in the regulation of the indoleamine-2,3-dioxygenase (IDO) activity in female blood monocytes has been studied in vitro. PSG stimulated IDO activity under the conditions of induction of the monocytes by interferon-γ. Upon the induction of cell proliferation by lipopolysaccharides, the stimulating effect was obtained only with 10 μg/mL of PSG. Enhanced IDO activity is probably a factor of peripheral immunological tolerance and antimicrobial protection against intracellular infections in the gestation period.     

Characterization of a Brassica napus myrosinase pseudogene: myrosinases are members of the BGA family of β-glycosidases

Myrosinase isoenzymes are known to be encoded by two different families of genes denoted MA and MB. Nucleotide sequence analysis of a Brassica napus genomic clone containing a gene for myrosinase revealed it to be a pseudogene of the MA family. The gene spans more than 5 kb and contains at least 12 exons. The exon sequence of the gene is highly similar to myrosinase cDNA sequences. However, the gene displays three potential or actual pseudogene characters. Southern blot analysis using probes from the 3 portions of the genomic and B. napus MA and MB cDNA clones showed that MA type myrosinases are encoded by approximately 4 genes, while MB type myrosinases are encoded by more than 10 genes in B. napus. Northern blots with mRNA from seeds and young leaves probed with the MA-and MB-specific probes showed that the MA and MB myrosinase gene families are differentially expressed. Myrosinases are highly similar to proteins of a -glycosidase enzyme family comprising both -glycosidases and phospho--glycosidases of as diverged species as archaebacteria, bacteria, mammals and plants. By homology to these -glycosidases, putative active site residues in myrosinase are discussed on the basis of the similarity between -glycosidases and cellulases.     

Evolutionary analysis of the multigene pregnancy-specific β1-Glycoprotein family: Separation of historical and nonhistorical signals

The pregnancy-specific ₁-glycoproteins (PSG) form a large family of closely related proteins. Using newly developed methods of sequence analysis, in combination with protein modeling, we provide a framework for investigating the evolution and biological function of genes like the PSG. Evolutionary trees, based on C-terminal sequence, group PSG genes in a manner consistent with their genomic organization. Trees constructed using the N-terminal domain sequences are unreliable as an indicator of phylogeny because of non-neutral processes of sequence change. During duplication of the PSG genes, evolutionary pressures have resulted in a gradient of constrained change across each gene. The N-terminal domains show a nonrandom pattern of amino acid substitutions clustered in the immunoglobulin complementarity-determining region (CDR)-like regions, which appear to be important in the function of the protein.     

A new rare phenotype of glycine-rich β-glycoprotein

Summary A rare phenotype of serum glycine-rich -glycoprotein was found in a healthy Swiss woman. The rare gene product was also observed in three of her four sons.     

Are the pH-sensitive mutants members of theVMA family?

This work was supported by Grant 7S 203 013 07.     

Further family studies on the genetic control of β2-glycoprotein I concentration in human serum

Summary 88 families with a total of 213 children were examined for ₂-glycoprotein I serum concentrations. In 74 families parents and children had normal concentrations. In 9 families one of the parents and approximately half of the children had intermediate concentrations. These individuals are presumably heterozygous for a deficiency gene Bg^D. In these families ₂-glycoprotein I concentration appears to be controlled by a pair of alleles which are transmitted as autosomal co-dominants. The results in 5 families did not conform to this genetic hypothesis, since children with an intermediate concentration of ₂-glycoprotein I were found whose both parents had a normal concentration of this protein. Non-genetic factors may be responsible for phenotypic variations in the different genetic types.Supported by U.S.P.H.S. Grant AM 11796-02 and by the Deutsche Forschungsgemeinschaft.     

Characterization of subfractions of β2-glycoprotein I: Evidence for sialic acid microheterogeneity

• 1.1. β₂-Glycoprotein I is a sialic acid microheterogeneous protein and contains on the average 11 mol sialic acid/mol.
• 2.2. Linear correlation was found between sialic acid content and pI of isolated subfractions.
• 3.3. Asialo-β₂2-glycoprotein I consists of 2 isoforms. Each of which can originate from the same subfraction.
• 4.4. The isolated subfractions exhibited almost the same amino acid composition.

     

Modulation of natural and interleukin-2-induced tumour-cytolytic activities by the members of a protein family related to β-thromboglobulin

Summary A preparation of three C-terminal fragments of the platelet protein -thromboglobulin was previously described to have immunomodulatory properties on phagocytic cells. One of the components is obviously identical to the recently described neutrophil-activating peptide 2 (NAP-2). In further investigations on this protein preparation (called factor C) we are able to show an additional influence on the tumour-cytolytic activities of mononuclear cells. Total neutralization of the factor C effect, by treating a factor C preparation with specific monoclonal antibody C24 prior to application in cell culture, proved that the effect is really restricted to factor C proteins. If factor C is given in combination with natural interleukin-2 (IL-2) a dose-dependent suppression of IL-2-mediated natural killer lymphokine-activated killer activity can be measured, which is first detectable 72 h after addition of factor C. Suppression does not occur if the both factors are added within a time interval of more than 12 h. Depletion of monocytes from mononuclear cells has no effect on factor-C-mediated cytotoxicity, demonstrating that factor C acts directly on lymphoid cells.     

Characterization of a new β-glucosidase/β-xylosidase from the gut microbiota of the termite (Reticulitermes santonensis)

The gut of the termite Reticulitermes santonensis contains an interesting diversity of prokaryotic and eukaryotic microorganisms not found elsewhere. These microorganisms produce many enzyme-digesting lignocellulosic compounds, probably in cooperation with endogenous enzymes. Regarding cellulose and hemicellulose digestion in the termite gut, much remains to be learned about the relative contributions of termite enzymes and enzymes produced by different microorganisms. Here we grew bacterial colonies from termite gut suspensions, identifying 11 of them after PCR amplification of their 16S rRNA genes. After constructing in Escherichia coli a genomic DNA library corresponding to all of the colonies obtained, we performed functional screening for α-amylase, xylanase, β-glucosidase, and endoglucanase activities. This screen revealed a clone producing β-glucosidase activity. Sequence analysis showed that the cloned genomic DNA fragment contained three complete ORFs (bglG, bglF, and bglB) organized in a putative bgl operon. The new β-glucosidase (BglB), identified with its regulators BglG and BglF, belongs to glycoside hydrolase family 1. The new β-glucosidase was expressed in E. coli and purified by affinity chromatography. The purified enzyme shows maximal activity at pH 6.0 and 40?°C. It also displays β-xylosidase activity.     

Identification of two Drosophila TGF-β family members in the grasshopper Schistocerca americana

Intercellular signaling molecules of the transforming growth factor- (TGF-) superfamily are required for pattern formation in many multicellular organisms. The decapentaplegic (dpp) gene of Drosophila melanogaster has several developmental roles. To improve our understanding of the evolutionary diversification of this large family we identified dpp in the grasshopper Schistocerca americana. S. americana diverged from D. melanogaster approximately 350 million years ago, utilizes a distinct developmental program, and has a 60-fold-larger genome than D. melanogaster. Our analyses indicate a single dpp locus in D. melanogaster and S. americana, suggesting that dpp copy number does not correlate with increasing genome size. Another TGF- superfamily member, the D. melanogaster gene 60A, is also present in only one copy in each species. Comparison of homologous sequences from D. melanogaster, S. americana, and H. sapiens, representing roughly 900 million years of evolutionary distance, reveals significant constraint on sequence divergence for both dpp and 60A. In the signaling portion of the dpp protein, the amino acid identity between these species exceeds 74%. Our results for the TGF- superfamily are consistent with current hypotheses describing gene duplication and diversification as a frequent response to high levels of selective pressure on individual family members.     

Characterization of β-galactosidases from the germinating seeds of Vigna sinensis

Four forms of β-galactosidase from the germinating seeds of Vigna sinensis were separated and partially purified by ammonium sulphate precipitation, ion exchange chromatography (DE-52) and gel filtration to more than 50% purity as judged by PAGE. The pH and temperature optima, stability, M_r, kinetic parameters and energy of activation of each enzyme have been determined. The four forms differed in their M,_s and ionic charges.     

Characterization of transgenic rice plants over-expressing the stress-inducible β-glucanase gene Gns1

The Gns1 gene of rice (Oryza sativa L. japonica) encodes 1,3;1,4- glucanase (EC 3.2.1.73), which hydrolyzes 1,3;1,4--glucosidic linkages on 1,3;1,4--glucan, an important component of cell walls in the Poaceae family. RNA and protein gel blot analyses demonstrated that blast disease or dark treatment induced the expression of the Gns1 gene. To assess the function of the Gns1 gene in disease resistance, we characterized transgenic rice plants constitutively expressing the Gns1 gene. The introduced Gns1 gene was driven by the CaMV 35S promoter and its products were found in the apoplast and accumulated in up to 0.1% of total soluble protein in leaves. Although transgenic plants showed stunted growth and impaired root formation, fertility, germination, and coleoptile elongation appeared unaffected compared to non-transgenic control plants, indicating that Gns1 does not play a crucial role in rice germination and coleoptile elongation. When transgenic plants were inoculated with virulent blast fungus (Magnaporthe grisea), they developed many resistant-type lesions on the inoculated leaf accompanying earlier activation of defense-related genes PR-1 and PBZ1 than in control plants. Transgenic plants spontaneously produced brown specks, similar in appearance to those reported for an initiation type of disease-lesion-mimic mutants, on the third and fourth leaves and occasionally on older leaves without inoculation of pathogens. Expression of the two defense-related genes was drastically increased after the emergence of the lesion-mimic phenotype.     

Co-stimulatory members of the TNFR family: keys to effective T-cell immunity?

Interactions between co-stimulatory ligands and their receptors are crucial for the activation of T cells, the prevention of tolerance and the development of T-cell immunity. It is now evident that members of the immunoglobulin-like CD28-B7 co-stimulatory family cannot fully account for an effective long-lasting T-cell response or the generation of memory T cells. Several members of the tumour-necrosis factor receptor (TNFR) superfamily--OX40, 4-1BB, CD27, CD30 and HVEM (herpes-virus entry mediator)--are poised to deliver co-stimulatory signals both early and late after encounter with antigen. The roles of these molecules in initiating and sustaining the T-cell response and in promoting long-lived immunity are discussed.     

Characterization of the α- and β-Mannosidases of Porphyromonas gingivalis

Mannose is an important sugar in the biology of the Gram-negative bacterium Porphyromonas gingivalis. It is a major component of the oligosaccharides attached to the Arg-gingipain cysteine proteases, the repeating units of an acidic lipopolysaccharide (A-LPS), and the core regions of both types of LPS produced by the organism (O-LPS and A-LPS) and a reported extracellular polysaccharide (EPS) isolated from spent culture medium. The organism occurs at inflamed sites in periodontal tissues, where it is exposed to host glycoproteins rich in mannose, which may be substrates for the acquisition of mannose by P. gingivalis. Five potential mannosidases were identified in the P. gingivalis W83 genome that may play a role in mannose acquisition. Four mannosidases were characterized in this study: PG0032 was a β-mannosidase, whereas PG0902 and PG1712 were capable of hydrolyzing p-nitrophenyl α-d-mannopyranoside. PG1711 and PG1712 were α-1→3 and α-1→2 mannosidases, respectively. No enzyme function could be assigned to PG0973. α-1→6 mannobiose was not hydrolyzed by P. gingivalis W50. EPS present in the culture supernatant was shown to be identical to yeast mannan and a component of the medium used for culturing P. gingivalis and was resistant to hydrolysis by mannosidases. Synthesis of O-LPS and A-LPS and glycosylation of the gingipains appeared to be unaffected in all mutants. Thus, α- and β-mannosidases of P. gingivalis are not involved in the harnessing of mannan/mannose from the growth medium for these biosynthetic processes. P. gingivalis grown in chemically defined medium devoid of carbohydrate showed reduced α-mannosidase activity (25%), suggesting these enzymes are environmentally regulated.