Small Angle X-ray Scattering-Based Elucidation of the Self-Association Mechanism of Human Insulin Analogue LysB29(N蔚蠅-carboxyheptadecanoyl) des(B30)

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• 作者：Malene Hillerup Jensen ; Per-Olof Wahlund ; Katrine N酶rgaard Toft ; Jes Kristian Jacobsen ; Dorte Bjerre Steensgaard ; Marco van de Weert ; Svend Havelund ; Bente Vestergaard
• 刊名：Biochemistry
• 出版年：2013
• 出版时间：January 15, 2013
• 年：2013
• 卷：52
• 期：2
• 页码：282-294
• 全文大小：491K
• 年卷期：v.52,no.2(January 15, 2013)
• ISSN：1520-4995

文摘

Lys^B29(N^蔚蠅-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 鈥渢ense鈥?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 鈥渞elaxed鈥?state R₆-hexamer and an R₃T₃T₃R₃-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.