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Biomedical subjects

C P Vogt

Publications and source records attributed to C P Vogt.

3 recordsLinked to original sources
PubMed1997

Metallothionein accretion in human hepatic cells is linked to cellular proliferation.

The basal amounts of metallothionein (MT) and its rates of biosynthesis were compared in resting and proliferating Chang liver (CCl-13) cells. In resting cells the total amounts of the detectable isoforms MT-2 and MT-1e were approx. 1.6x10(6) and 4x10(5) molecules per cell respectively. In exponentially growing cultures the cellular contents of both isoforms increased co-ordinately approx. 4-fold and decreased again to the initial values within 48 h after entering density-mediated growth arrest. As documented for MT-2 its transient accretion was attributable to a 10-fold rise in the rate of biosynthesis of this protein during the growth phase. Measurements of the relative amounts of MT-2 mRNA indicated the occurrence of a more than 50% increase within the first 12 h after subculturing of the cells, followed by a return to basal levels thereafter. These results denote a direct link between the programming of MT synthesis and proliferation and thus attest to a central housekeeping function of the MTs.

Blotting, Northern↗
PubMed1996

Identification of stress proteins in lysates of human cell lines separated by two-dimensional electrophoresis and electroblotted simultaneously onto two different membranes.

A protocol based on a combination of established methods for the characterization and identification of inducible stress proteins in human cell lines is described. A particular protein spot, collected from several micropreparative two-dimensional electrophoresis (2-DE) gels, is concentrated into a new gel prior to simultaneous electrotransfer onto a Cationic Durapore (CD) membrane and onto a polyvinylidene (PVDF) backup membrane. The protein blotted onto the PVDF support is subjected to N-terminal sequence analysis. From the protein bound to the CD membrane the peptide mass profile is obtained by proteolytic digestion of the protein followed by the separation of the resulting peptides by high performance liquid chromatography (HPLC) and their detection by on-line electrospray mass spectrometry (LC/MS). Additional internal sequence information may be obtained by amino acid sequence analysis of peptides collected in the HPLC effluent. The efficiency of this strategy is demonstrated with two proteins extracted from 15 micropreparative 2-DE gels of an extract of a human liver cell line. The peptide mass fingerprinting of a 60 kDa protein with a pI of 5.3 assigned 22 of 37 peptides to the heat shock protein 60 (Hsp 60). The result was confirmed by the N-terminal sequence analysis of the undigested protein and of an internal tryptic fragment. The second sample, a 40 kDa protein with a pI of 4.9, was identified as a processed form of the heat shock cognate 71 kDa protein (Hsc 70).

Amino Acid Sequence↗