Hormonal Properties of Avidin-Biotinylinsulin and Avidin-Biotinylcorticotropin Complexes

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Abstract
In connection with the development of affinity columns (based on the avidin-biotin interaction) for retrieval of peptide and protein hormone receptor, the hormonal properties of a number of avidin-biotinylinsulin and avidin-biotinylcorticotropin complexes were examined. Of particular interest was an evaluation of streptavidin as a ligand for the attachment of biotinylated hormones to solid supports and its possible advantage over SpHPP-avidin (S = succinoylated; pHPP = 3-(p-hydroxyphenyl)propionyl). As concerns binding kinetics using rat liver plasma membranes, streptavidin was superior to avidin since it does not display apparently nonsaturable binding. Scatchard analyses of the binding of 125I-streptavidin, 125I-S-streptavidin and 125I-SpHPP-avidin to rat liver plasma membranes gave Kd values of 6.7, 13.2 and 10.6 nM, respectively. The binding was saturable and the unlabeled proteins competed with their labeled counterparts for the membrane binding sites. Biotinylinsulin, attached to either streptavidin or SpHPP-avidin was able to compete for 125I-insulin-binding sites on rat liver plasma membranes though somewhat larger concentrations of the complexes than of insulin were required to achieve comparable inhibition. The median inhibitory dose values for insulin and the biotinylinsulin complexes were 5 and 80 nM, respectively. Biotinylcorticotropin was a more effective activator of particulate rat adrenal adenylate cyclase when complexed with unmodified avidin than with streptavidin, S-streptavidin or SpHPP-avidin.

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