Use of a urea and guanidine-HCl—propanol solvents system to purify a growth inhibitory glycopeptide by high-performance liquid chromatography

Sharifi, Behrooz G.; Bascom, Charles C.; Khurana, Vijay K.; Johnson, Terry C.

doi:10.1016/s0021-9673(01)81316-9

Cited by 21 publications

(8 citation statements)

References 19 publications

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“…Purification of the SGP The SGP inhibitor was released from intact bovine cerebral cortex cells by mild proteolysis, and purified to apparent homogeneity as previously described (Sharifi et al, 1985(Sharifi et al, ,1986a. Briefly, bovine cerebral cortex cells were incubated with dilute pronase for 15 minutes, after which the macromolecules in the supernatant fluid were ethanol precipitated, chloroformimethanol extracted (2:17 volivol), and further purified by DEAE ion-exchange chromatography, lectin affinity chroma-tography, and HPLC gel filtration chromatography.…”

Section: Methodsmentioning

confidence: 99%

Effects of a sialoglycopeptide on early events associated with signal transduction

Toole-Simms

Loder

Fattaey

et al. 1991

Journal Cellular Physiology

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A sialoglycopeptide (SGP), isolated and purified from bovine cerebral cortex cells, was studied in regard to early signal transduction events associated with the cell cycle. Previously shown to be a potent antagonist to a variety of mitogens, the SGP abrogated the ability of 12-O-tetradecanoylphorbol-13 acetate (TPA) to elicit an alkalinization of 3T3 cell cytosol, but only when added minutes prior to, or simultaneously with, the tumor promoter. 3T3 cell TPA-mediated Ca2+ mobilization was also inhibited by the SGP although the inhibitor itself did not bind Ca2+ in a cell-free assay. The results are discussed in light of the already known kinetics of interaction between the SGP, various mitogens, and the calcium ionophore A23187 with regard to the pivotal events leading to the decision of a cell to divide or not to divide.

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Section: Methodsmentioning

confidence: 99%

Effects of a sialoglycopeptide on early events associated with signal transduction

Toole-Simms

Loder

Fattaey

et al. 1991

Journal Cellular Physiology

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“…The SGP, which is a hydrophilic fragment of a larger glycoprotein (Sharifi et al, 1985), has a molecular weight of 18,000 and a PI of 3.0, and the biological activity appears to reside in the polypeptide sequence of the molecule (Sharifi et al, 198613). There are specific, high-affinity receptors on the surface of 3T3 cells for the SGP, and the inhibition of protein synthesis is correlated with the occupancy of the receptor .…”

mentioning

confidence: 99%

“…The isolation and purification to homogeneity of our sialoglycopeptide (SGP) inhibitor obtained from intact bovine cerebral cortex cells have been described previ-ously (Sharifi et al, 1986a;Bascom et al, 1986). The SGP, which is a hydrophilic fragment of a larger glycoprotein (Sharifi et al, 1985), has a molecular weight of 18,000 and a PI of 3.0, and the biological activity appears to reside in the polypeptide sequence of the molecule (Sharifi et al, 198613). There are specific, high-affinity receptors on the surface of 3T3 cells for the SGP, and the inhibition of protein synthesis is correlated with the occupancy of the receptor .…”

mentioning

confidence: 99%

Inhibition of DNA synthesis and cell division by a cell surface sialoglycopeptide

Fattaey

Johnson

Chou

1989

Journal Cellular Physiology

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We have isolated and purified a cell surface sialoglycopeptide (SGP) from bovine cerebral cortex cells that previously was shown to be a potent inhibitor of cellular protein synthesis. The following studies were carried out to characterize the potential ability of the SGP to inhibit DNA synthesis and to arrest cell division. Treatment of exponentially proliferating Swiss 3T3 cells with the SGP inhibitor resulted in a marked inhibition of thymidine incorporation within 24 h. When the SGP was removed from inhibited cultures, a sharp rise in 3H-thymidine incorporation followed within 3-4 h that peaked well above that measured in exponentially growing cultures, suggesting that the inhibitory action of the SGP was reversible and that a significant proportion of the arrested cells was synchronized in the mitotic cycle. In addition to DNA synthesis, the inhibitory action of the SGP was monitored by direct measurement of cell number. Consistent with the thymidine incorporation data, the SGP completely inhibited 3T3 cell division 20 h after its addition to exponentially growing cultures. Upon reversal there was a delay of 15 h before cell division resumed, when the arrested cells quickly doubled. Most, if not all, of the growth-arrested cells appeared to have been synchronized by the SGP. The SGP inhibited DNA synthesis in a surprisingly wide variety of target cells, and the relative degree of their sensitivity to the inhibitor was remarkably similar. Cells sensitive to the SGP ranged from vertebrate to invertebrate cells, fibroblast and epitheliallike cells, primary cells and established cell cultures, as well as a wide range of transformed cell lines.

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“…The peak resolution is excellent and about 98% purification can be achieved in one step. In the case of a hydrophobic protein, the yield can be improved further by incorporating suitable denaturant in the mobile phase [114]. Gelfiltration -size exclusion chromatography is often used to purify protein in the denatured state.…”

Section: Purification Of Protein In the Denatured Statementioning

confidence: 99%

Inclusion bodies and purification of proteins in biologically active forms

Mukhopadhyay

1997

Biotreatment, Downstream Processing and Modelling

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Use of a urea and guanidine-HCl—propanol solvents system to purify a growth inhibitory glycopeptide by high-performance liquid chromatography

Cited by 21 publications

References 19 publications

Effects of a sialoglycopeptide on early events associated with signal transduction

Effects of a sialoglycopeptide on early events associated with signal transduction

Inhibition of DNA synthesis and cell division by a cell surface sialoglycopeptide

Inclusion bodies and purification of proteins in biologically active forms

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