Exon 11 enhances insulin binding affinity and tyrosine kinase activity of the human insulin proreceptor.
Data presented here show that there are significant differences in the insulin binding affinity and the tyrosine kinase activity of the insulin proreceptor isoforms which contain or lack exon 11. The exon 11(+) proreceptor does not show significant variations from the mature processed insulin receptor and has only a 3- to 4-fold reduced affinity for insulin. In contrast, the exon 11(-) proreceptor showed a markedly reduced insulin binding (25- to 50-fold less) when assayed on intact cells. Upon solubilization of the cells, exon 11(-) proreceptor bound insulin with somewhat higher affinity. Mild trypsin treatment of the cells expressing either isoform of the insulin proreceptor restored insulin binding to near-normal levels. Analysis of tyrosine kinase activity revealed that the exon 11(+) proreceptor required somewhat higher concentrations of insulin than the mature processed receptor to achieve maximal autophosphorylation. The exon 11(-) proreceptor failed to fully phosphorylate even at 10(-6) M insulin. Thus, the presence or absence of this short sequence of 12 amino acids affects the folding and/or conformation of the proreceptor so as to confer altered binding of insulin. We suggest that in the absence of exon 11 the proreceptor assumes a strained conformation that disrupts the insulin binding site. Cleavage of the proreceptor at the alpha-beta-subunit junction then allows the alpha-subunit to achieve its normal binding conformation.
Pashmforoush
M
M
Department of Biochemistry and Molecular Biology, University of Chicago, Illinois 60637.
Yoshimasa
Y
Y
Steiner
D F
DF
D000345:Affinity Labels; D002478:Cells, Cultured; D003432:Cross-Linking Reagents; D005091:Exons; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D011498:Protein Precursors / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011972:Receptor, Insulin / Q000378:metabolism*
D000345:Affinity Labels; D003432:Cross-Linking Reagents; D007328:Insulin; D011498:Protein Precursors; C051512:insulin proreceptor; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin