| 1. |
- Fernanda Troncoso, M., et al.
(författare)
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A novel trypsin inhibitor from Peltophorum dubium seeds, with lectin-like properties, triggers rat lymphoma cell apoptosis
- 2003
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Ingår i: Archives of Biochemistry and Biophysics. - 0003-9861 .- 1096-0384. ; 411:1, s. 93-104
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Tidskriftsartikel (refereegranskat)abstract
- A trypsin inhibitor (PDTI) was isolated from Peltophorum dubium seeds by affinity chromatography on a thyroglobulin-agarose or a trypsin-agarose column. In both cases, SDS-PAGE showed two bands of M(r) 20,000 and 22,000, which could not be resolved. Their amino-terminal sequences were identical and similar to that of Kunitz-type soybean trypsin inhibitor (SBTI). Mass spectrometry analysis of tryptic digests of both bands showed 16 coincident peaks, suggesting that they are closely related proteins. The K(i)s for trypsin and chymotrypsin inhibitory activity of PDTI were 1.6 x 10(-7) and 1.3 x 10(-5)M, respectively. Lectin-like activity of PDTI and SBTI, detected by hemagglutination of rabbit erythrocytes, was inhibited by sialic acid-containing compounds. PDTI and SBTI caused apoptosis of Nb2 rat lymphoma cells, demonstrated by decrease of viability, DNA hypodiploidy, DNA fragmentation, and caspase-3-like activity. They had no effect on normal mouse splenocytes or lymphocytes, whereas they caused apoptosis of concanavalin A-stimulated mouse lymphocytes.
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| 2. |
- Pirpignani, María L., et al.
(författare)
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Structural and immunological aspects of Polybia scutellaris Antigen 5
- 2002
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Ingår i: Archives of Biochemistry and Biophysics. - 0003-9861 .- 1096-0384. ; 407:2, s. 224-230
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Tidskriftsartikel (refereegranskat)abstract
- Vespid venoms contain Antigen 5, an important allergen whose primary structure and immunological behavior have been extensively studied from venoms of vespids of the Northern Hemisphere. We report herein structural and immunological aspects of Antigen 5 from Polybia scutellaris subspecies rioplatensis (vulgar name: camoati) found in South America. Mast cell degranulation, histamine release, and IgE induction experiments performed in mice allow us to suggest that P. scutellaris Antigen 5 is a variant with reduced IgE response and anaphylactic activity. Sequence data indicate that the protein has a 72.5-90.3% similarity to that of members of the vespid Antigen 5 family with an already known primary structure. Moreover, results suggest that the protein-a new member of an extracellular protein superfamily-could be a good candidate for immunotherapy related to vespid allergy.
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| 3. |
- Stark, Katarina, et al.
(författare)
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Expression of CYP4F8 (prostaglandin H 19-hydroxylase) in human epithelia and prominent induction in epidermis of psoriatic lesions
- 2003
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Ingår i: Archives of Biochemistry and Biophysics. - 0003-9861 .- 1096-0384. ; 409:1, s. 188-196
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Tidskriftsartikel (refereegranskat)abstract
- Our aim was to determine the tissue distribution of CYP4F8, which occurs in human seminal vesicles and catalyzes 19-hydroxylation of prostaglandin H(1) and H(2) in vitro (J. Bylund, M. Hidestrard, M. Ingelman-Sundberg, E.H. Oliw, J. Biol. Chem. 275 (2000) 21844-21849). Polyclonal antibodies were raised in rabbits against RVEPLG, the C-terminal end of CYP4F8, and purified by affinity chromatography. Screening of 50 human tissues for CYP4F8 immunoreactivity revealed protein expression, inter alia, in seminal vesicles, epidermis, hair follicles, sweat glands, corneal epithelium, proximal renal tubules, and epithelial linings of the gut and urinary tract. The CYP4F8 transcripts were detected by reverse transcription polymerase chain reaction and by Northern blot analysis. There was a prominent induction of CYP4F8 immunoreactivity and mRNA in psoriasis in comparison with unaffected epidermis of the same patients. The cDNA of CYP4F8 from plucked scalp hair roots was identical with the genital cDNA sequence. We conclude that CYP4F8 is present in epithelial linings and up regulated in epidermis of psoriatic lesions.
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| 5. |
- Zhao, Ming, 1966-, et al.
(författare)
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Vascular smooth muscle cell proliferation requires both p38 and BMK1 MAP kinases
- 2002
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Ingår i: Archives of Biochemistry and Biophysics. - 0003-9861 .- 1096-0384. ; 400:2, s. 199-207
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Tidskriftsartikel (refereegranskat)abstract
- Vascular smooth muscle cell (VSMC) proliferation is a key event in the progression of atherosclerosis. Induction of both c-fos (through the transcription factor Elk-1) and c-jun, both immediate early genes, is important for the stimulation of VSMC proliferation and migration. It was earlier found that p38 mitogen-activated protein (MAP) kinase upregulates c-jun gene transcription through phosphorylation of two myocyte enhancer factor 2 (MEF2) family transcription factors, MEF2A and MEF2C, while big MAP kinase 1 (BMK1) may upregulate c-jun gene transcription through MEF2A, MEF2C, and also MEF2D. Here, we report that inhibition of BMK1 by a dominant negative form of MEK5 or pharmacologic inhibition of p38 by SB 203580 additively suppress serum-induced VSMC proliferation. This additive effect of p38 and BMK1 inhibition implies that these two kinases coordinately regulate MEF2 transcription factors. The exclusive activation of MEF2D by BMK1 appears required for this cooperative upregulation of c-jun in VSMC, and coactivation of p38 and BMK1 also has additive effects on the activation of a reporter gene linked to the c-jun promoter in our experimental system. Thus, coordinate activity of both the p38 and BMK1 pathways appears necessary for optimal transcription of c-jun and, pari pasu, VSMC proliferation. These results may have implications for the future design of pharmacologic agents for inhibition of VSMC growth.
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| 6. |
- Buss, Joan L, et al.
(författare)
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Oxidative stress mediates toxicity of pyridoxal isonicotinoyl hydrazone analogs
- 2004
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Ingår i: Archives of Biochemistry and Biophysics. - : Elsevier BV. - 0003-9861 .- 1096-0384. ; 421:1
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Tidskriftsartikel (refereegranskat)abstract
- Pyridoxal isonicotinoyl hydrazone (PIH) and many of its analogs are effective iron chelators in vivo and in vitro, and are of interest for the treatment of secondary iron overload. Because previous work has implicated the Fe3+-chelator complexes as a determinant of toxicity, the role of iron-based oxidative stress in the toxicity of PIH analogs was assessed. The Fe3+ complexes of PIH analogs were reduced by K562 cells and the physiological reductant, ascorbate. Depletion of the antioxidant, glutathione, sensitized Jurkat T lymphocytes to the toxicity of PIH analogs and their Fe 3+ complexes, and toxicity of the chelators increased with oxygen tension. Fe3+ complexes of pyridoxal benzoyl hydrazone (PBH) and salicyloyl isonicotinoyl hydrazone (SIH) caused lipid peroxidation and toxicity in K562 cells loaded with eicosapentenoic acid (EPA), a readily oxidized fatty acid, whereas Fe(PIH)2 did not. The lipophilic antioxidant, vitamin E, completely prevented both the toxicity and lipid peroxidation caused by Fe(PBH)2 in EPA-loaded cells, indicating a causal relationship between oxidative stress and toxicity. PBH also caused concomitant lipid peroxidation and toxicity in EPA-loaded cells, both of which were reversed as its concentration increased. In contrast, PIH was inactive, while SIH was equally toxic toward control and EPA-loaded cells, without causing lipid peroxidation, indicating a much smaller contribution of oxidative stress to the mechanism of toxicity of these analogs. In summary, PIH analogs and their Fe3+ complexes are redox active in the intracellular environment. The contribution of oxidative stress to the overall mechanism of toxicity varies across the series. © 2003 Elsevier Inc. All rights reserved.
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| 7. |
- Cisneros, David A., et al.
(författare)
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Inversion of the allosteric response of Escherichia coli glucosamine-6-P deaminase to N-acetylglucosamine 6-P, by single amino acid replacements
- 2004
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Ingår i: Archives of Biochemistry and Biophysics. - : Elsevier BV. - 0003-9861 .- 1096-0384. ; 421:1, s. 77-84
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Tidskriftsartikel (refereegranskat)abstract
- Amino acid replacements in the active site of glucosamine-6-P deaminase from Escherichia coli (GlcN6P deaminase, EC 3.5.99.6) involving the residues D141 and E148 produce atypical allosteric kinetics. These residues are located in the chain segment 139-156 which is part of the active site and which also forms several intersubunit contacts close to the allosteric site. In the D141N and E148Q mutant forms of this deaminase, there is an inversion of the effect of its physiological allosteric effector, N-acetylglucosamine 6-P, which becomes an inhibitor at substrate concentrations above a critical value. For both mutants, this particular point appears at low substrate concentration and the inhibition by the allosteric activator is the dominant effect in velocity versus substrate curves. These effects are analyzed as a particular case of the concerted allosteric model, assuming that the R state, the conformer displaying the higher affinity for the substrate, is the less catalytic state, thus producing an inverted allosteric response.
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| 8. |
- Jortikka, Matti, et al.
(författare)
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The role of microtubules in the regulation of proteoglycan synthesis in chondrocytes under hydrostatic pressure.
- 2000
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Ingår i: Archives of Biochemistry and Biophysics. - : Elsevier. - 0003-9861 .- 1096-0384. ; 374:2, s. 172-180
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Tidskriftsartikel (refereegranskat)abstract
- Chondrocytes of the articular cartilage sense mechanical factors associated with joint loading, such as hydrostatic pressure, and maintain the homeostasis of the extracellular matrix by regulating the metabolism of proteoglycans (PGs) and collagens. Intermittent hydrostatic pressure stimulates, while continuous high hydrostatic pressure inhibits, the biosynthesis of PGs. High continuous hydrostatic pressure also changes the structure of cytoskeleton and Golgi complex in cultured chondrocytes. Using microtubule (MT)-affecting drugs nocodazole and taxol as tools we examined whether MTs are involved in the regulation of PG synthesis in pressurized primary chondrocyte monolayer cultures. Disruption of the microtubular array by nocodazole inhibited [(35)S]sulfate incorporation by 39-48%, while MT stabilization by taxol caused maximally a 17% inhibition. Continuous hydrostatic pressure further decreased the synthesis by 34-42% in nocodazole-treated cultures. This suggests that high pressure exerts its inhibitory effect through mechanisms independent of MTs. On the other hand, nocodazole and taxol both prevented the stimulation of PG synthesis by cyclic 0. 5 Hz, 5 MPa hydrostatic pressure. The drugs did not affect the structural and functional properties of the PGs, and none of the treatments significantly affected cell viability, as indicated by the high level of PG synthesis 24-48 h after the release of drugs and/or high hydrostatic pressure. Our data on two-dimensional chondrocyte cultures indicate that inhibition of PG synthesis by continuous high hydrostatic pressure does not interfere with the MT-dependent vesicle traffic, while the stimulation of synthesis by cyclic pressure does not occur if the dynamic nature of MTs is disturbed by nocodazole. Similar phenomena may operate in cartilage matrix embedded chondrocytes.
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| 9. |
- Landberg, Eva, 1966-, et al.
(författare)
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Changes in Glycosylation of Human Bile-Salt-Stimulated Lipase during Lactation
- 2000
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Ingår i: Archives of Biochemistry and Biophysics. - : Elsevier BV. - 0003-9861 .- 1096-0384. ; 377:2, s. 246-254
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Tidskriftsartikel (refereegranskat)abstract
- Bile-salt-stimulated lipase (BSSL) is an enzyme in human milk, which is important for the fat digestion in the newborn infant. BSSL is highly glycosylated and includes one site for N-glycosylation and several sites for O-glycosylation. BSSL has previously been found to express Lewis a, Lewis b, and Lewis x carbohydrate antigens. In this study, glycosylation of BSSL was studied at different times during lactation. BSSL was purified from milk collected individually from four donors at several different times during the first 6 months of lactation. The BSSL glycans were characterized through monosaccharide analysis, high-pH anion-exchange chromatography, matrix-assisted laser desorption–ionization mass spectrometry, and ELISA. Both total carbohydrate content and relative amount of sialic acid were higher in BSSL from the first lactation month as compared to BSSL from milk collected later in lactation. BSSL from the first lactation month also showed a different composition of sialylated O-linked glycans and the N-linked oligosaccharides consisted of lower amounts of fucosylated structures compared to later in lactation. We also found a gradual increase in the expression of the carbohydrate epitope Lewis x on BSSL throughout the lactation period. This study shows that glycosylation of BSSL is dependent on blood group phenotype of the donor and changes substantially during the lactation period.
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| 10. |
- Påhlsson, Peter, 1962-, et al.
(författare)
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Characterization of galactosyl glycerolipids in the HT29 human colon carcinoma cell line
- 2001
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Ingår i: Archives of Biochemistry and Biophysics. - : Elsevier BV. - 0003-9861 .- 1096-0384. ; 396:2, s. 187-198
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Tidskriftsartikel (refereegranskat)abstract
- Glycoglycerolipids constitute a family of glycolipids with apparently very restricted expression in human tissues. They have previously been detected only in the testis and the nervous system. In the present study, two glycoglycerolipids were isolated from the HT29 human colon carcinoma cell line. The glycoglycerolipids were structurally characterized as a monogalactosylglycerolipid (1-O-alkyl-2-O-acyl-3-O-(▀-galactosyl)-sn-glycerol) and a digalactosylglycerolipid (1-O-alkyl-2-O-acyl-3-O-(▀-galactosyl(1-4)a-galactosyl)-sn- glycerol) using NMR and mass spectrometry. This digalactosylglycerolipid has not previously been structurally characterized. When HT29 cells were allowed to differentiate into more enterocyte-like cells by culture in glucose-free medium, expression of both of these glycoglycerolipids was greatly diminished. The presence of glycoglycerolipids in a human colon carcinoma cell line indicates that expression of this family of glycolipids may not be as restricted as previously thought. Instead this class of glycolipids may serve as differentiation antigens in various normal tissues and in tumor development. The Gala1-4Gal epitope was previously identified as a receptor for bacterial adhesins and toxins. The finding that this epitope is also linked to a glycerolipid moiety opens up new possible roles for this carbohydrate receptor in intracellular signaling.
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| 11. |
- Öllinger, Karin, 1962-
(författare)
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Inhibition of cathepsin D prevents free-radical-induced apoptosis in rat cardiomyocytes
- 2000
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Ingår i: Archives of Biochemistry and Biophysics. - : Elsevier BV. - 0003-9861 .- 1096-0384. ; 373:2, s. 346-351
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Tidskriftsartikel (refereegranskat)abstract
- Apoptosis was inhibited in rat cardiomyocytes pretreated with the aspartic protease inhibitor pepstatin A and subsequently exposed to naphthazarin (5,8-dihydroxy-1,4-naphthoquinone). Cathepsin D was released from lysosomes to the cytosol upon exposure to naphthazarin, and the enzyme activity decreased simultaneously. Later, cathepsin D reappeared in granules of increased size, and enzyme activity was restored. Activation of caspase-3- like proteases was detected, and the number of cells showing apoptotic morphology increased with time. Pepstatin A pretreatment did not prevent release of cathepsin D from lysosomes but did significantly inhibit subsequent naphthazarin-induced caspase activation and apoptotic morphology. This suggests that cathepsin D exerts its apoptosis-stimulating effect upstream of caspase-3-like activation. (C) 2000 Academic Press.
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| 21. |
- Eklund, Erik, et al.
(författare)
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Dermatan is a better substrate for 4-O-sulfation than chondroitin : implications in the generation of 4-O-sulfated, L-iduronate-rich galactosaminoglycans
- 2000
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Ingår i: Archives of Biochemistry and Biophysics. - : Elsevier BV. - 0003-9861. ; 383:2, s. 171-177
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Tidskriftsartikel (refereegranskat)abstract
- The biosynthesis of dermatan sulfate is a complex process that involves, inter alia, formation of L-iduronic acid residues by C5-epimerization of D-glucuronic acid residues already incorporated into the growing polymer. It has been shown previously that this reaction is promoted by the presence of the sulfate donor 3'-phosphoadenosine-5'-phosphosulfate. In the present investigation, the role of sulfation in the biosynthesis of L-iduronic acid-rich galactosaminoglycans was examined more closely by a study of the substrate specificities and kinetic properties of the sulfotransferases involved in dermatan sulfate biosynthesis. Comparison of the acceptor reactivities of oligosaccharides from chondroitin and dermatan, in an in vitro system containing microsomes from cultured human skin fibroblasts and 3'-phosphoadenosine-5'-phosphosulfate, showed that Km values for the dermatan fragments were substantially lower than those for their chondroitin counterparts. Calculation of Vmax values likewise showed that dermatan was the better substrate. Whereas dermatan incorporated [35S]sulfate exclusively at the C4 position of N-acetylgalactosamine residues, approximately equal amounts of radioactivity were found at the C4 and C6 positions in the labelled chondroitin. Under standard assay conditions, the 4-O-sulfation of dermatan proceeded about six times faster than the 4-O-sulfation of chondroitin. On the basis of these results, we propose that L-iduronic acids, formed in the course of the biosynthesis of dermatan sulfates, enhance sulfation of their adjacent N-acetylgalactosamine residues, and will thereby be locked in the L-ido configuration.
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| 22. |
- Falkenberg, Cecilia, et al.
(författare)
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Expression of a functional proteinase inhibitor capable of accepting xylose: bikunin
- 2001
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Ingår i: Archives of Biochemistry and Biophysics. - : Elsevier BV. - 0003-9861. ; 387:1, s. 99-106
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Tidskriftsartikel (refereegranskat)abstract
- Bikunin is a Kunitz-type proteinase inhibitor, which is cross-linked to heavy chains via a chondroitin sulfate chain, forming inter-alpha-inhibitor and related molecules. Rat bikunin was produced by baculovirus-infected insect cells. The protein could be purified with a total yield of 20 mg/liter medium. Unlike naturally occuring bikunin the recombinant protein had no galactosaminoglycan chain. Endoglycosidase digestion also suggested that the recombinant form lacked N-linked oligosaccharides. Bikunin is translated as a part of a precursor, alpha1-microglobulin/bikunin, but the functional significance of the cotranslation is unknown. Our results indicate that the proteinase inhibitory function of bikunin is not regulated by the alpha1-microglobulin-part of the alpha1-microglobulin/bikunin precursor since recombinant bikunin had the same trypsin inhibitory activity as the recombinant precursor. Both free bikunin and the precursor were also functional as a substrate in an in vitro xylosylation system. This demonstrates that the alpha1-microglobulin-part is not necessary for the first step of galactosaminoglycan assembly.
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| 35. |
- Sternberg, Lawrence R, et al.
(författare)
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Alternative splicing of the human MUC2 gene.
- 2004
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Ingår i: Archives of biochemistry and biophysics. - : Elsevier BV. - 0003-9861. ; 421:1, s. 21-33
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Tidskriftsartikel (refereegranskat)abstract
- Human colon cancers differ in amounts of MUC2 mucin synthesized. However, it is unclear whether MUC2 encodes a single protein. When clones of human colon cancer cells were assayed with antibodies against the TR2 mucin repeat or non-TR2 epitopes, differences in relative expression of MUC2 proteins suggested multiple immunoreactive forms. RT-PCR analysis detected the established 15kbp MUC2 cDNA and a novel form (designated MUC2.1) lacking the MUC2 TR2 repeat. Sequencing of cDNA and genomic DNA indicated that MUC2.1 results from an alternate splice donor. RT-PCR with splice-junction spanning primers confirmed the expression of MUC2.1 mRNA. Anti-MUC2.1 antibody stained colon cancer cells and normal colon in a pattern different from TR2-specific antibody. The presence of MUC2.1 mucin may help us to explain previous conflicting reports that have attempted to correlate the relative abundance of MUC2 protein and/or mRNA with the biological behavior of colon cancer cells.
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