Insulin bound

To explain the complex of phenomena observed in Fig. 2.5, we postulate that insulin is present in the subcutaneous depot in three different forms free insulin on dimeric form, free insulin on hexameric form, and insulin bound to various substances and surfaces in the tissue. Moreover, we assume that only dimeric insulin can diffuse across the capillary wall. In qualitative terms, our explanation is then ... 

G Free FTTC-Insulin Bound FtTC-insulin [1 A A. /1... 

Brownlee M and Cerami A. A glucose-controUed insuhn-delivery system Semisynthetic insulin bound to lectin. Science 1979 206 1190-1191. 

Our consideration of the signal-transduction cascades initiated by epinephrine and insulin included examples of how components of signal-transduction pathways are poised for action, ready to be activated by minor modifications. For example, G-protein a subunits require only the binding of GTP in exchange for GDP to transmit a signal. This exchange reaction is thermodynamically favorable, but it is quite slow in the absence of an appropriate activated 7TM receptor. Similarly, the tyrosine kinase domains of the dimeric insulin receptor are ready for phosphorylation and activation but require the presence of insulin bound between two a subunits to draw the activation loop of one tyrosine kinase into the active site of a partner tyrosine kinase to initiate this process. 

Liu, F., et al. Glucose-induced release of glycosylpoly(ethylene glycol) insulin bound to a soluble conjugate of concanavalin A. Bioconjugate Chemistry, 1997, 8(5), 664-672. 

All these findings suggested that insulin binding is not an artifact but reflects a true physicochemical combination between muscle cells and insulin. This conclusion found further support from experiments made by Stetten with radioactive insulin ( S- and labeled). The use of labeled insulin permitted Stetten to demonstrate that the insulin concentration is much higher within the tissues than in the medium, and the amount of insulin bound per unit mass was found to be the same when the hormone was injected as when it was added to the incubation mixture. After measuring the amount of bound insulin, the investigators could express the metabolic activity due to insulin—namely, glycogen synthesis per unit mass of bound insulin. These studies established that hormonal activity is proportional to the amount of hormone bound to the muscle cell. Hormonal activity was expressed per unit mass for the first time in these experiments. 

Brownlee, M., Cerami, A. (1979). Glucose-controlled insulin-delivery system - semisynthetic insulin bound to lectin. Science, 206,1190-1191. 

It would be highly desirable to check this point with radioactive insulin, by determining the localization of radioactive insulin bound to diaphragm by radioautography. 

The very same principle was applied by Suzuki et al. [19] for the purification of the placental insulin receptor, starting with a prepurified sample, obtained from a plasma membrane fraction of human placenta by a fractionation protocol including affinity chromatography on Sepharose-concanavalin A as the last step. The following purification step was carried out, using insulin bound to dextran (M 40000) as the biospecific, water-soluble carrier. After filtration of the dextran-insulin-receptor complex on an adequate gel and then dissociation by mild acidification, the insulin receptor was obtained in a purified form. This method permitted a 8700 fold purification, from crude plasma membrane and, even more remarkable, a 60-fold purification after the affinity chromatography step. 

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