N‐terminal amino acid sequences of precursor and mature forms of α‐1‐antitrypsin

Groß, Volker; Kaiser, Clemens; Tran‐Thi, Thuy‐Anh; Schmelzer, Elmon; Witt, I.; Plummer, Thomas H.; Heinrich, Peter C.

doi:10.1016/0014-5793(83)80069-6

Cited by 17 publications

(8 citation statements)

References 24 publications

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“…A 1 ml portion of medium or of the supernatant obtained from the cell homogenate was added to S ml of 20 mM-Tris/HCl, pH 7.5,0.14 M-NaCl, 5 mM-EDTA and 1 % Triton X-100. After addition of 10 ,ul of a specific antiserum against rat al-antitrypsin (Gross et al, 1983), rat a,-acid glyco-protein (Gross et al, 1984), human al-antitrypsin (Gross etal., 1990b) or human interleukin-6 (Gross et al, 1989) and incubation at 0°C overnight, the antigen-antibody complexes were bound to 10 mg (dry weight) of Protein A-Sepharose 4B and washed four times with the above-mentioned buffer and twice with 50 mM-sodium phosphate buffer, pH 7.5. Elution was performed by incubation with 0.1M-Tris/HCl (pH 6.8)/5 / , -mercaptoethanol/5 % SDS/ 10% glycerol at 95 'C for 5 min.…”

Section: Immunoprecipitation and Electrophoresismentioning

confidence: 99%

Inhibition of protein N-glycosylation by 2-deoxy-2-fluoro-d-galactose

Groß

Hull

Berger

et al. 1992

Biochemical Journal

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The effects of 2-deoxy-2-fluoro-D-galactose (dGalF) on N- and O-glycosylation of proteins was studied in rat hepatocyte primary cultures and in human monocytes. In hepatocytes, dGalF at concentrations of 1 mM or higher completely inhibited N-glycosylation of alpha 1-antitrypsin and alpha 1-acid glycoprotein, whereas 4 mM-2-deoxy-D-galactose (dGal) only slightly impaired N-glycosylation. In monocytes, 1 mM- or 4 mM-dGalF blocked N-glycosylation of alpha 1-antitrypsin and of interleukin-6, while O-glycosylation of interleukin-6 remained unaffected. In monocytes, dGal had no effect on protein N-glycosylation. Addition of uridine effectively prevented the UTP deficiency induced by dGalF, but had no effect on the inhibition of protein N-glycosylation by dGalF. Using 19F-n.m.r. spectroscopy, 2-deoxy-2-fluoro-D-galactose 1-phosphate (dGalF-1-P), UDP-dGalF and UDP-dGlcF could be identified as the major metabolites of dGalF in hepatocytes as well as in monocytes. In conclusion, compared with dGal, dGalF is a more efficient inhibitor of protein N-glycosylation. The effect is not caused by the depletion of UTP induced by dGalF, but rather by metabolites of dGalF. dGalF is metabolized not only in hepatocytes but also in peripheral blood monocytes, which can be used for ex vivo studies of disturbances in D-galactose metabolism.

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Section: Immunoprecipitation and Electrophoresismentioning

confidence: 99%

Inhibition of protein N-glycosylation by 2-deoxy-2-fluoro-d-galactose

Groß

Hull

Berger

et al. 1992

Biochemical Journal

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“…For this, 1.5-2.5 ml of medium or 0.4-0.8 ml of the supernatant obtained from the cell homogenate were added to 5 ml of 20 mM-Tris/HCl (pH 7.6)/0.14 M-NaCl/5 mM-EDTA/1 % Triton X-100 containing 1 mMphenylmethanesulphonyl fluoride and 0.1 mg of kallikrein trypsin inhibitor (kindly provided by Bayer A. G., Wuppertal-Elberfeld). After addition of 7.5 ,l of a specific antiserum against rat ax1PI (Gross et al, 1983c) or rat a,AGP (Gross et al, 1984) and incubation at 0°C overnight, the antigen-antibody complexes were bound to 7 mg (dry wt.) of protein A-Sepharose and washed four times with the above-mentioned buffer and twice with 50 mM-sodium phosphate buffer, pH 7.5.…”

Section: Immunoprecipitationmentioning

confidence: 99%

Different effects of the glucosidase inhibitors 1-deoxynojirimycin, N-methyl-1-deoxynojirimycin and castanospermine on the glycosylation of rat α1-proteinase inhibitor and α1-acid glycoprotein

Groß¹,

Tran‐Thi²,

Schwarz³

et al. 1986

Biochemical Journal

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The glucosidase inhibitors 1-deoxynojirimycin, N-methyl-1-deoxynojirimycin and castanospermine were used to inhibit oligosaccharide processing in primary cultures of rat hepatocytes. Their effect on the glycosylation of alpha 1-proteinase inhibitor (alpha 1PI) and alpha 1-acid glycoprotein (alpha 1AGP) was studied. Of the three glucosidase inhibitors examined, 1-deoxynojirimycin inhibited not only oligosaccharide trimming but also glycosylation de novo of newly synthesized proteins, resulting in the formation of alpha 1PI with two and three (normally carrying three) and alpha 1AGP with two to five (normally carrying six) oligosaccharide side chains. In the presence of the glucosidase inhibitors, glucosylated high-mannose-type oligosaccharides accumulated. Whereas most of the endoglucosaminidase-H-sensitive oligosaccharides formed in the presence of 1-deoxynojirimycin contained only one glucose residue, N-methyl-1-deoxynojirimycin and castanospermine led mainly to the formation of oligosaccharides with three glucose residues. None of the three glucosidase inhibitors completely prevented the formation of complex-type oligosaccharides. Thus, in their presence, alpha 1PI and alpha 1AGP with a mixture of both high-mannose and complex-type oligosaccharides were secreted.

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“…A1AT is a glycoprotein exhibiting extensive glycosylation following synthesis in epithelial cells such as hepatocytes [33]. Whereas unglycosylated A1AT migrates at *45 kDa size [34][35][36], we noted that ASCsecreted A1AT migrates at 54 kDa (Fig. 1D), similar to A1AT secreted by bronchial epithelial cells (Fig.…”

Section: Resultsmentioning

confidence: 57%

Oncostatin M and TNF-α Induce Alpha-1 Antitrypsin Production in Undifferentiated Adipose Stromal Cells

Mian

Meador

et al. 2017

Stem Cells and Development

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Alpha-1 antitrypsin (A1AT), a circulating acute-phase reactant antiprotease, is produced and secreted by cells of endodermal epithelial origin, primarily hepatocytes, and by immune cells. Deficiency of A1AT is associated with increased risk of excessive lung inflammation and injury, especially following chronic cigarette smoke (CS) exposure. Exogenous administration of mesenchymal progenitor cells, including adipose tissue-derived stromal/stem cells (ASC), alleviates CS-induced lung injury through paracrine effectors such as growth factors. It is unknown, however, if mesodermal ASC can secrete functional A1AT and if CS exposure affects their A1AT production. Human ASC collected via liposuction from nonsmoking or smoking donors were stimulated by inflammatory cytokines tumor necrosis alpha (TNFα), oncostatin M (OSM), and/or dexamethasone (DEX) or were exposed to sublethal concentrations of ambient air control or CS extract (0.5%-2%). We detected minimal expression and secretion of A1AT by cultured ASC during unstimulated conditions, which significantly increased following stimulation with TNFα or OSM. Furthermore, TNFα and OSM synergistically enhanced A1AT expression and secretion, which were further increased by DEX. The A1AT transcript variant produced by stimulated ASC resembled that produced by bronchial epithelial cells rather than the variant produced by monocytes/macrophages. While the cigarette smoking status of the ASC donor had no measurable effect on the ability of ASC to induce A1AT expression, active exposure to CS extract markedly reduced A1AT expression and secretion by cultured ASC, as well as human tracheobronchial epithelial cells. ASC-secreted A1AT covalently complexed with neutrophil elastase in control ASC, but not in cells transfected with A1AT siRNA. Undifferentiated ASC may require priming to secrete functional A1AT, a potent antiprotease that may be relevant to stem cell therapeutic effects.

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N‐terminal amino acid sequences of precursor and mature forms of α‐1‐antitrypsin

Cited by 17 publications

References 24 publications

Inhibition of protein N-glycosylation by 2-deoxy-2-fluoro-d-galactose

Inhibition of protein N-glycosylation by 2-deoxy-2-fluoro-d-galactose

Different effects of the glucosidase inhibitors 1-deoxynojirimycin, N-methyl-1-deoxynojirimycin and castanospermine on the glycosylation of rat α1-proteinase inhibitor and α1-acid glycoprotein

Oncostatin M and TNF-α Induce Alpha-1 Antitrypsin Production in Undifferentiated Adipose Stromal Cells

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