The insulin like growth factor (IGF) system is essential for the regulation of metabolic and mitogenic responses in mammalian cells. This system is regulated by three ligands, IGF-I, IGF-II and insulin, each sharing high degree of sequence and structural homology. The IGF system has been heavily implicated in promoting cancer progression, with several studies linking the upregulation of IGF-II, the insulin receptor isoform-A and IGF-1R with cancer growth and survival.
Currently, there is no structure of the entire IR-A or IGF-1R in complex with IGF-II. Mutagenesis of IGF-II defined two surfaces important for receptor binding (site 1 and 2). However, recent cryo electron microscopy (cryo-EM) structures of insulin and IGF-I complexed to their cognate receptors highlight a discrepancy, where all but one of the site 2 residues defined by mutagenesis are not making contact with the receptor.
Here we aim to use Fluorescence Resonance Energy Transfer (FRET) as a molecular ruler to measure the distance between the receptor and ligand site 2 residues.
We will report the recombinant expression, purification and refold of several IGF-II analogues for FRET analysis. The significance of site 2 to our study is that our preliminary structure/function data suggests this interaction site is involved in the growth promoting functions. To effectively inhibit cancer growth induced by IGF-II binding we need to further map the interactions using a different approach to precisely define the site 2 interaction. Understanding of the molecular mechanisms of activating growth and division will ultimately lead to the development of IGF-II analogues with antagonistic properties for the treatment of cancer.