TY - JOUR
T1 - Insulin and IGF binding by IGFBP-3 fragments derived from proteolysis, baculovirus expression and normal human urine
AU - Vorwerk, Peter
AU - Yamanaka, Yoshitaka
AU - Spagnoli, Anna
AU - Oh, Youngman
AU - Rosenfeld, Ron G.
PY - 1998
Y1 - 1998
N2 - Recombinant human IGFBP-3 was proteolysed with different concentrations of plasmin for various periods of time. The major IGFBP-3 fragment resulting from this digestion migrated at ca. 15 kDa in nonreducing SDS-PAGE. Following the identification of this fragment as an N-terminal IGFBP-3 fragment, by use of N-terminus-specific monoclonal antibody and amino acid sequence analysis, we constructed and expressed a similar fragment in a baculovims expression system. The fragments resulting from plasmin digestion, as well as the baculovirus-expressed recombinant human IGFBP-31-97, retain weak IGF binding and show specific insulin binding on cross-linking and western ligand blot. RhIGFBP-31-97 can inhibit insulin receptor autophosphorylation in insulin receptor-overexpressing NIH 3T3 cells. Insulin and IGF binding to IGFBP-3 fragments could be further demonstrated in normal urine. These data indicate the physiological significance of IGFBP-3 fragments derived from proteolysis in vivo.
AB - Recombinant human IGFBP-3 was proteolysed with different concentrations of plasmin for various periods of time. The major IGFBP-3 fragment resulting from this digestion migrated at ca. 15 kDa in nonreducing SDS-PAGE. Following the identification of this fragment as an N-terminal IGFBP-3 fragment, by use of N-terminus-specific monoclonal antibody and amino acid sequence analysis, we constructed and expressed a similar fragment in a baculovims expression system. The fragments resulting from plasmin digestion, as well as the baculovirus-expressed recombinant human IGFBP-31-97, retain weak IGF binding and show specific insulin binding on cross-linking and western ligand blot. RhIGFBP-31-97 can inhibit insulin receptor autophosphorylation in insulin receptor-overexpressing NIH 3T3 cells. Insulin and IGF binding to IGFBP-3 fragments could be further demonstrated in normal urine. These data indicate the physiological significance of IGFBP-3 fragments derived from proteolysis in vivo.
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U2 - 10.1210/jcem.83.4.4858
DO - 10.1210/jcem.83.4.4858
M3 - Article
C2 - 9543173
AN - SCOPUS:0031762905
SN - 0021-972X
VL - 83
SP - 1392
EP - 1395
JO - Journal of Clinical Endocrinology and Metabolism
JF - Journal of Clinical Endocrinology and Metabolism
IS - 4
ER -