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  • 1995-1999  (2)
  • 1980-1984
  • 1950-1954
  • 1999  (2)
  • 1982
  • 1952
  • 2Cys-peroxiredoxin  (1)
  • HPLC  (1)
  • 1
    Electronic Resource
    Electronic Resource
    Isolation, Characterization, and Stability of Positional Isomers of Mono-PEGylated Salmon Calcitonins (1999)
    Springer
    Pharmaceutical research 16 (1999), S. 813-818 
    Details
    ISSN: 1573-904X
    Keywords: calcitonin ; polyethylene glycol ; PEGylation ; peptide ; tryptic digestion ; stability ; HPLC
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract Purpose. To separate and characterize the different positional isomers of mono-PEGylated salmon calcitonins (mono-PEG-sCTs) and to evaluate the effects of the PEGylation site on the stability of different mono-PEG-sCTs in rat kidney homogenate. Methods. Mono-PEG-sCTs were prepared using succinimidyl carbonate monomethoxy polyethylene glycol (5,000 Da) and separated by gel-filtration HPLC followed by reversed-phase HPLC. To characterize PEGylated sCTs, matrix-assisted laser desorption ionization time of flight mass spectrometry (M ALDI-TOF MS) and reversed-phase HPLC of the trypsin digested samples were performed. Mono-PEG-sCTs and sCT in rat kidney homogenates were measured by column-switching reversed-phase HPLC with on-line detection of the radioiodinated samples using a flow-through radioisotope detector. Results. Three different mono-PEGylated sCTs were separated by reversed-phase gradient HPLC. From the MALDI-TOF MS analysis, the average molecular weight of mono-PEG-sCTs was confirmed as around 8650 Da. The presence of PEG moiety in the mono-PEG-sCTs was also manifested by the fact that the distance between two adjacent mass spectum lines was 44 Da which corresponds to PEG monomer unit. Tryptic digestion analysis demonstrated that these mono-PEG-sCTs are 3 positional isomers of N-terminus, Lys18- and Lys11-residue modified mono-PEGylated sCTs. The degradation half-life of these 3 positional isomers in rat kidney homogenates significantly increased in order of the N-terminus (125.5 min), Lys11- (157.3 min), and Lysl8-residue modified mono-PEGylated sCT (281.5 min) over the native sCT (4.8 min). Conclusions. Three positional isomers of mono-PEGylated sCTs were purified and characterized. Of these, the resistance to proteolytic degradation was highest for the Lysl8-residue modified mono-PEG-sCT. These studies demonstrate that the in vivo stability of PEGylated sCTs is highly dependent on the site of PEG molecule attachment.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1018861616465
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    Paper (German National Licenses)
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  • 2
    Electronic Resource
    Electronic Resource
    Molecular cloning, expression, and functional characterization of a 2Cys-peroxiredoxin in Chinese cabbage (1999)
    Springer
    Plant molecular biology 40 (1999), S. 825-834 
    Details
    ISSN: 1573-5028
    Keywords: 2Cys-peroxiredoxin ; Chinese cabbage ; expression ; functional characterization ; gene cloning
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A cDNA (C2C-Prx) corresponding to a 2Cys-peroxiredoxin (2Cys-Prx) was isolated from a leaf cDNA library of Chinese cabbage. The predicted amino acid sequence of C2C-Prx has 2 conserved cysteines and several peptide domains present in most of the 2Cys-Prx subfamily members. It shows the highest sequence homology to the 2Cys-Prx enzymes of spinach (88%) and Arabidopsis (86%). Southern analysis using the cDNA insert of C2C-Prx revealed that it consists of a small multigene family in Chinese cabbage genome. RNA blot analysis showed that the gene was predominantly expressed in the leaf tissue of Chinese cabbage seedlings, but the mRNA was generally expressed in most tissues of mature plant, except roots. The expression of C2C-Prx was slightly induced by treatment with H2O2 (100μM) or Fe3+/O2/DTT oxidation system, but not by ABA (50 μM) or GA3 (10 μM). The C2C-Prx is encoded as a preprotein of 273 amino acids containing a putative chloroplast-targeting signal of 65 amino acids at its N-terminus. The N-terminally truncated recombinant protein (ΔC2C-Prx) migrates as a dimer in a non-reducing SDS-polyacrylamide gel and as a monomer in a reducing condition. The ΔC2C-Prx shows no immuno cross-reactivity to antiserum of the yeast thiol-specific antioxidant protein, and vice versa. The ΔC2C-Prx prevents the inactivation of glutamine synthetase and the DNA cleavage in the metal-catalyzed oxidation system. In the yeast thioredoxin system containing thioredoxin reductase, thioredoxin, and NADPH, the ΔC2C-Prx exhibits peroxidase activity on H2O2.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006271823973
    Library Location Call Number Volume/Issue/Year Availability
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    Paper (German National Licenses)
    Fulltext
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