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81.
Berit M. Mortensen Hanne W. Lund Greg Jablonski Ruth H. Paulssen Jan O. Gordeladze 《Bioscience reports》1995,15(3):135-150
In normal rats treated with 1,25(OH)2D3 or 24,25(OH)2D3, serum Ca2+, ALP, PRL and GH are significantly altered. In order to study the primary effect of vitamin D3 analogues on target organ function, rat UMR 106 osteosarcoma and GH3 pituitary adenoma cells in monolayer culture were exposed accordingly.Surprisingly, prolonged exposure of these cell lines to physiological levels of either 1,25(OH)2D3 or 24,25(OH)2D3 did not significantly affect the secretory parameters (ALP, PRL or GH) tested. However, 1,25(OH)2D3 exposure significantly reduced PTH- and Gpp(NH)p-elicited AC as well as Gpp(NH)p-stimulated PLC activities in the UMR 106 cells. These changes were accompanied by an increase and decrease in the membrane contents of the G-protein subunits G36 and Gq/11, respectively. In contrast, 24,25(OH)2D3 remained without significant biological effect on these signalling systems despite concomitantly augmented levels of G36. TRH- and Gpp(NH)p-elicited PLC activities in the GH3 cells were significantly reduced by 1,25(OH)2D3 with a concurrent reduction in cellular amounts of Gq/11, however, 24,25(OH)2D3 did not significantly alter any signalling systems nor G-proteins analyzed.It is concluded that the osteoblastic and pituitary cell secretion of ALP, PRL and GH remain unaffected by the presence of 1,25(OH)2D3 and 24,25(OH)2D3, despite distinct alterations in components of G-protein mediated signalling pathways. Hence, other factors like ambient Ca2+ may be responsible for the perturbed secretory patterns of ALP and PRL seen in vitamin D3 treated rats.Abbreviations AC
adenylate cyclase
- ALP
alkaline phosphatase
- BGP
osteocalcin
- BSA
bovine serum albumin
- DA
dopamine
- DAG
diacylglycerol
- GH
growth hormone
- GHRH
growth hormone releasing hormone
- Gpp(NH)p
guanosine 5-[-imido]triphosphate
- G-protein
guanine nucleotide-binding regulatory protein
- Gs etc.
Gs protein -subunit
- IP3
inositol 1,4,5 trisphosphate
- OAF
osteoclast activating factor
- PGE2
prostaglandin E2
- PKA & PKC
protein kinase A & C
- PLC
phospholipase C
- PRL
prolactin
- PTH
parathyroid hormone
- SRIF
somatostatin
- TRH
thyrotropin releasing hormone
- VIP
vasoactive intestinal peptide
- 25(OH)D3
25 hydroxy vitamin D3
- 1,25(OH)2D3
1·25 dihydroxy vitamin D3
- 24,25(OH)2D3
24,25 dihydroxy vitamin D3 相似文献
82.
83.
High level biosynthesis and secretion of the thermostable hybrid (1-3,1-4)--glucanase H(A16-M) has been achieved inSaccharomyces cerevisiae by means of the yeast vacuolar endoprotease B promoter (PRB1p) and theBacillus macerans (1-3,1-4)--glucanase signal peptide. The N-glycans present on the yeast-secreted H(A16-M), denoted H(A16-M)-Y, were released by endoglycosidase H, and identified by proton NMR spectroscopy to be a homologous series of Man8-13GlcNAc2, although only traces of Man9GlcNAc2 were found. Therefore, processing of N-glycans on H(A16-M)-Y is similar to that on homologous proteins. Most of the N-glycans (88%) were neutral while the remainder were charged due to phosphorylation. Site-directed mutagenesis of Asn to Gln in two of the N-glycosylation sequons, and subsequent analysis of the N-glycans on the yeast-secreted proteins together with analysis of the N-glycans from the individual sites of H(A16-M)-Y suggest the presence of steric hindrance to glycan modification by the glycans themselves. H(A16-M)-Y produced under control of either the yeast protease B or the yeast 3-phosphoglycerate kinase promoter, each in two differentSaccharomyces strains revealed a dependence of N-glycan profile on both strain and culture conditions. The extent of O-glycosylation was found to be nine mannose units per H(A16-M)-Y molecule. An attempt to identify the linkage-sites for the O-glycans by amino acid sequencing failed, suggesting non-stoichiometric or heterogeneous O-glycosylation. The possible modes in which N-glycans might contribute to resistance of H(A16-M)-Y to irreversible thermal denaturation are discussed with respect to structural information available for H(A16-M)-Y.
Abbreviations: AMY,B. amyloliquefaciens (1-3,1-4)--glucanse; MAC,B. macerans (1-3,1-4)--glucanase H(A16-M), H(A36-M), H(A78-M),H(A107-M) and H(A152-M), hybrid (1-3,1-4)--glucanases containing 16, 36, 78, 107 and 152 N-terminal amino acids, respectively, derived from AMY with the remaining amino acids derived from MAC; similar enzyme abbreviations followed by Y, e.g. H(A16-M)-Y, denote the enzymes secreted from yeast cells; PCR, polymerase chain reaction; PGKp, yeast 3-phosphoglycerate kinase promoter; PRB1p, yeast protease B promoter; LB, Luria-Bertani medium; SC, minimal medium; CNBr, cyanogen bromide; Endo Hf, endoglycosidase H fusion protein; PNGase F, peptide:N-glycosidase F; HPAEC; high pH anion exchange chromatography; HVE, high voltage paper electrophoresis; CPY, yeast carboxypeptidase Y. 相似文献
84.
We present kinetic studies on the enzymatic transfer of several synthetic sialic acid analogues, modified at C-5, to distinct glycoprotein glycans by sialytransferases differing in acceptor- and linkage-specificity. Biochemical properties of sialic acids were modified by introducing formyl-, trifluoroacetyl-, benzyloxycarbonyl-, and aminoacetyl-groups to the amino group at C-5 of neuraminic acid. The latter substitution renders the corresponding -glyocoside resistant towards sialidases. The respective CMP-sialic acid analogues were prepared by CMP-sialic acid synthase with a yield of 13–55%.The kinetic parameters of several sialyltransferases for the 5-substituted CMP-glycosides differed significantly. Relative to parent CMP-NeuAc, reaction rates of human- and rat liver Gal1, 4GlcNAc 2,6-sialyl-transferases ranged from 50 to 170%, of GalNAc 2,6-sialyltransferases from 40–140%, and of Gal1,3Gal-NAc 2,3-sialyltransferase from 20–50%. Resialylation of asialo-1-acid glycoprotein by 5-N-formyl- and 5-N-aminoacetyl-neuraminic acid employing rat liver Gal1,4GlcNAc 2,6-sialyltransferase proceeded to about 80% of galactose sites which is identical to the extent achieved with parent NeuAc.According to our data, neosialoglycoconjugates which carry sialic acids modified at theN-acetyl group can be prepared for structure-function analysis, as this position seems crucial for recognition of adhesion proteins and influenza viruses. 相似文献
85.
86.
O. Basso J. Olsen A. M. Johansen K. Christensen 《BMJ (Clinical research ed.)》1997,315(7121):1498-1502
OBJECTIVE: To estimate the risk of having a low birthweight infant associated with changes in social, environmental, and genetic factors. DESIGN: Population based, historical cohort study using the Danish medical birth registry and Statistic Denmark''s fertility database. SUBJECTS: All women who had a low birthweight infant (< 2500 g) (index birth) and a subsequent liveborn infant (outcome birth) in Denmark between 1980 and 1992 (exposed cohort, n = 11,069) and a random sample of the population who gave birth to an infant weighing > or = 2500 g and to a subsequent liveborn infant (unexposed cohort, n = 10,211). MAIN OUTCOME MEASURES: Risk of having a low birthweight infant in the outcome birth as a function of changes in male partner, area of residence, type of job, and social status between the two births. RESULTS: Women in the exposed cohort showed a high risk (18.5%) of having a subsequent low birthweight infant while women in the unexposed cohort had a risk of 2.8%. After adjustment for initial social status, a decline in social status increased the absolute risk of having a low birthweight infant by about 5% in both cohorts, though this was significant only in the unexposed cohort. Change of male partner did not modify the risk of low birth weight in either cohort. CONCLUSION: Having had a low birthweight infant and a decline in social status are strong risk factors for having a low birthweight infant subsequently. 相似文献
87.
Søren Bak Rachel Alice Kahn Carl Erik Olsen Barbara Ann Halkier 《The Plant journal : for cell and molecular biology》1997,11(2):191-201
Obtusifoliol 14β-demethylase from Sorghum bicolor (L.) Moench has been cloned using a gene-specific probe generated using PCR primers designed from an internal 14 amino acid sequence. The sequence identifies sorghum obtusifoliol 14α-demethylase as a cytochrome P450 and it is assigned to the CYP51 family together with the sterol 14α-demethylases from fungi and mammals. The presence of highly conserved regions in the amino acid sequences, analogous substrates and the same metabolic role demonstrate that the sterol 14α-demethylases are orthologous enzymes. The sterol 14α-demethylases catalyse an essential step in sterol biosynthesis as evidenced by the absence of a 14α-methyl group in all known functional sterols. A functional sorghum obtusifoliol 14α-demethylase was expressed at high levels in Escherichia coli and purified using an efficient method based on temperature-induced Triton X-114 phase partitioning. The recombinant purified enzyme produced a type I spectrum with obtusifoliol as substrate. Reconstitution of purified recombinant enzyme with sorghum NADPH—cytochrome P450 reductase in dilaurylphosphatidylcholine micelles confirms that obtusifoliol 14α-demethylase catalyses the 14α-demethylation of obtusifoliol to 4α-methyl-5α-ergosta-8,14,24(28)-trien-3β-ol as evidenced by GC—MS. The isolation of a cDNA clone encoding the plant sterol 14α-demethylase, combined with the previously isolated cDNA clones for fungal and mammalian sterol 14α-demethylases, provides an important tool in the rational design of specific inhibitors towards the individual sterol 14α-demethylases. 相似文献
88.
Akira F. Peters Madeleine J. H. van Oppen Christian Wiencke Wytze T. Stam Jeanine L. Olsen 《Journal of phycology》1997,33(2):294-309
Phylogenetic relationships in the Desmarestiales (Phaeophyceae) were inferred among the monotypic Arthrocladia (Arthrocladiaceae) and 27 isolates from Desmarestiaceae, representing 17 taxa of Desmarestia and the monotypic Antarctic genera Himantothallus and Phaeurus. Phaeurus and Arthrocladia were used as outgroups. Parsimony analyses of nuclear ribosomal DNA internal transcribed spacer (ITS1 and ITS2) sequences, in which gaps were both included and excluded, yielded well-resolved trees with a consistent general branching pattern. A parallel analysis of nine morphological and life-history characters and three ecological characters yielded a similar tree but provided little resolution in the terminal clades. The position of the monotypic Arthrocladia villosa within the Desmarestiales is consistent with monophyly for the order, but its position as the most primitive desmarestialean is not resolvable from the molecular data set. The basal position of Phaeurus, the Antarctic Desmarestia species, and Himantothallus is consistent with the hypothesis of a Southern Hemisphere origin for the family Desmarestiaceae. The more recent Northern Hemisphere “aculeata” clade evolved from an Antarctic ancestor. A “D. aculeata-like” species was ancestral to a lineage characterized by annual sporophytes with high sulfuric acid content, which radiated into many species, widely distributed in both hemispheres. Mapping of morphological and ecological characters onto the molecular tree confirm the informativeness of sulfuric acid-containing vacuoles and unilocular sporangial types. There is good congruence between phylogenetic tree topology and temperature impints in relation to biogeographic distribution, supporting the theory that temperature tolerance is a conservative trait. 相似文献
89.
Nucleotide sequence analysis of genes encoding a toluene/benzene-2-monooxygenase from Pseudomonas sp. strain JS150. 总被引:7,自引:5,他引:2
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It was previously shown by others that Pseudomonas sp. strain JS150 metabolizes benzene and alkyl- and chloro-substituted benzenes by using dioxygenase-initiated pathways coupled with multiple downstream metabolic pathways to accommodate catechol metabolism. By cloning genes encoding benzene-degradative enzymes, we found that strain JS150 also carries genes for a toluene/benzene-2-monooxygenase. The gene cluster encoding a 2-monooxygenase and its cognate regulator was cloned from a plasmid carried by strain JS150. Oxygen (18O2) incorporation experiments using Pseudomonas aeruginosa strains that carried the cloned genes confirmed that toluene hydroxylation was catalyzed through an authentic monooxygenase reaction to yield ortho-cresol. Regions encoding the toluene-2-monooxygenase and regulatory gene product were localized in two regions of the cloned fragment. The nucleotide sequence of the toluene/benzene-2-monooxygenase locus was determined. Analysis of this sequence revealed six open reading frames that were then designated tbmA, tbmB, tbmC, tbmD, tbmE, and tbmF. The deduced amino acid sequences for these genes showed the presence of motifs similar to well-conserved functional domains of multicomponent oxygenases. This analysis allowed the tentative identification of two terminal oxygenase subunits (TbmB and TbmD) and an electron transport protein (TbmF) for the monooxygenase enzyme. In addition to these gene products, all the tbm polypeptides shared significant homology with protein components from other bacterial multicomponent monooxygenases. Overall, the tbm gene products shared greater similarity with polypeptides from the phenol hydroxylases of Pseudomonas putida CF600, P35X, and BH than with those from the toluene monooxygenases of Pseudomonas mendocina KR1 and Burkholderia (Pseudomonas) pickettii PKO1. The relationship found between the phenol hydroxylases and a toluene-2-monooxygenase, characterized in this study for the first time at the nucleotide sequence level, suggested that DNA probes used for surveys of environmental populations should be carefully selected to reflect DNA sequences corresponding to the metabolic pathway of interest. 相似文献
90.