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

TY - JOUR

T1 - Small angle X-ray scattering-based elucidation of the self-association mechanism of human insulin analogue lys(B29)(N(e)¿-carboxyheptadecanoyl) des(B30)

AU - Jensen, Malene Hillerup

AU - Wahlund, Per-Olof

AU - Toft, Katrine Nørgaard

AU - Jacobsen, Jes Kristian

AU - Steensgaard, Dorte Bjerre

AU - van de Weert, Marco

AU - Havelund, Svend

AU - Vestergaard, Bente

PY - 2013/1/15

Y1 - 2013/1/15

N2 - LysB29(Nεω-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 (T6-hexamers) as the smallest repeating unit. The smallest oligomer building block in the process is a T6T6-dihexamer. This tense dihexamer is formed by the allosteric change of the initial equilibrium between a proposed "relaxed" state R6-hexamer and an R3T 3T3R3-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.

AB - LysB29(Nεω-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 (T6-hexamers) as the smallest repeating unit. The smallest oligomer building block in the process is a T6T6-dihexamer. This tense dihexamer is formed by the allosteric change of the initial equilibrium between a proposed "relaxed" state R6-hexamer and an R3T 3T3R3-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.

U2 - 10.1021/bi3008615

DO - 10.1021/bi3008615

M3 - Journal article

C2 - 23256662

SN - 0006-2960

VL - 52

SP - 282

EP - 294

JO - Biochemistry

JF - Biochemistry

IS - 2

ER -