Small angle X-ray scattering-based elucidation of the self-association mechanism of human insulin analogue lys(B29)(N(e)¿-carboxyheptadecanoyl) des(B30)

Malene Hillerup Jensen, Per-Olof Wahlund, Katrine Nørgaard Toft, Jes Kristian Jacobsen, Dorte Bjerre Steensgaard, Marco van de Weert, Svend Havelund, Bente Vestergaard

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15 Citations (Scopus)

Abstract

Lys(B29)(N(e)¿-carboxyheptadecanoyl) des(B30) human insulin is an insulin analogue belonging to a class of analogues designed to form soluble depots in subcutis by self-association, aiming at a protracted action. On the basis of small angle X-ray scattering (SAXS) supplemented by a range of biophysical and structural methods (field flow fractionation, dynamic and multiangle light scattering, circular dichroism, size exclusion chromatography, and crystallography), we propose a mechanism for the self-association expected to occur upon subcutaneous injection of this insulin analogue. SAXS data provide evidence of the in solution structure of the self-associated oligomer, which is a long straight rod composed of "tense" state insulin hexamers (T(6)-hexamers) as the smallest repeating unit. The smallest oligomer building block in the process is a T(6)T(6)-dihexamer. This tense dihexamer is formed by the allosteric change of the initial equilibrium between a proposed "relaxed" state R(6)-hexamer and an R(3)T(3)T(3)R(3)-dihexamer. The allosteric change from relaxed to tense is triggered by removal of phenol, mimicking subcutaneous injection. The data hence provide the first unequivocal evidence of the mechanism of self-association for this type of insulin analogue.
Original languageEnglish
JournalBiochemistry
Volume52
Issue number2
Pages (from-to)282-94
Number of pages13
ISSN0006-2960
DOIs
Publication statusPublished - 15 Jan 2013

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