Protein Engineering, Vol. 12, No. 4, 297-303,
April 1999
© 1999 Oxford University Press
Modelling of the disulphide-swapped isomer of human insulin-like growth factor-1: implications for receptor binding
1 Department of Biochemistry, School of Biological Sciences, University of Southampton, 6 Bassett Crescent East, Southampton SO16 7PX, 2 Department of Chemistry, University of York, Heslington, York YO1 5DD, UK, 3 Hagedorn Research Institute, Niels Steensens Vej 6, DK-2820 Gentofte, Denmark, 4 Department of Crystallography, Birkbeck College, Malet Street, London WC1E 7HX, UK and 5 Institut für Biochemie, Rheinisch-Westfälische Technische Hochschule Aachen, Klinikum, Pauwelsstrasse 30, D-52057 Aachen, Germany
Insulin-like growth factor-1 (IGF-1) is a serum protein which unexpectedly folds to yield two stable tertiary structures with different disulphide connectivities; native IGF-1 [18–61,6–48,47–52] and IGF-1 swap [18–61,6–47, 48–52]. Here we demonstrate in detail the biological properties of recombinant human native IGF-1 and IGF-1 swap secreted from Saccharomyces cerevisiae. IGF-1 swap had a ~30 fold loss in affinity for the IGF-1 receptor overexpressed on BHK cells compared with native IGF-1.The parallel increase in dose required to induce negative cooperativity together with the parallel loss in mitogenicity in NIH 3T3 cells implies that disruption of the IGF-1 receptor binding interaction rather than restriction of a post-binding conformational change is responsible for the reduction in biological activity of IGF-1 swap. Interestingly, the affinity of IGF-1 swap for the insulin receptor was ~200 fold lower than that of native IGF-1 indicating that the binding surface complementary to the insulin receptor (or the ability to attain it) is disturbed to a greater extent than that to the IGF-1 receptor. A 1.0 ns high-temperature molecular dynamics study of the local energy landscape of IGF-1 swap resulted in uncoiling of the first A-region 
-helix and a rearrangement in the relative orientation of the A- and B-regions. The model of IGF-1 swap is structurally homologous to the NMR structure of insulin swap and CD spectra consistent with the model are presented. However, in the model of IGF-1 swap the C-region has filled the space where the first A-region -helix has uncoiled and this may be hindering interaction of Val44 with the second insulin receptor binding pocket.
Keywords: insulin/insulin-like growth factor-1/molecular dynamics/receptor binding/structure–function relationships
6 To whom correspondence should be addressed. E-mail, steve{at}soton.ac.uk
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