Todorova Nevena, Zlatic Courtney O, Mao Yu, Yarovsky Irene, Howlett Geoffrey J, Gooley Paul R, Griffin Michael D W
School of Engineering, RMIT University , GPO Box 2476V, Melbourne, Victoria 3001, Australia.
Department of Biochemistry and Molecular Biology, University of Melbourne , Parkville, Victoria 3010, Australia.
Biochemistry. 2017 Mar 28;56(12):1757-1767. doi: 10.1021/acs.biochem.6b01146. Epub 2017 Mar 15.
The apolipoprotein family is structurally defined by amphipathic α-helical regions that interact with lipid surfaces. In the absence of lipid, human apolipoprotein (apo) C-II also forms well-defined amyloid fibrils with cross-β structure. Formation of this β-structure is accompanied by the burial of two charged residues, K30 and D69, that form an ion-pair within the amyloid fibril core. Molecular dynamics (MD) simulations indicate these buried residues form both intra- and intersubunit ion-pair interactions that stabilize the fibril. Mutations of the ion-pair (either K30D or D69K) reduce fibril stability and prevent fibril formation by K30D apoC-II under standard conditions. We investigated whether mixing K30D apoC-II with other mutants would overcome this loss of fibril forming ability. Co-incubation of equimolar mixtures of K30D apoC-II with wild-type, D69K, or double-mutant (K30D/D69K) apoC-II promoted the incorporation of K30D apoC-II into hybrid fibrils with increased stability. MD simulations showed an increase in the number of intersubunit ion-pair interactions accompanied the increased stability of the hybrid fibrils. These results demonstrate the important role of both intra- and intersubunit charge interactions in stabilizing apoC-II amyloid fibrils, a process that may be a key factor in determining the general ability of proteins to form amyloid fibrils.
载脂蛋白家族在结构上由与脂质表面相互作用的两亲性α-螺旋区域定义。在没有脂质的情况下，人载脂蛋白（apo）C-II也会形成具有交叉β结构的明确淀粉样纤维。这种β结构的形成伴随着两个带电残基K30和D69的埋藏，它们在淀粉样纤维核心内形成离子对。分子动力学（MD）模拟表明，这些埋藏的残基形成亚基内和亚基间的离子对相互作用，从而稳定纤维。离子对的突变（K30D或D69K）会降低纤维稳定性，并在标准条件下阻止K30D apoC-II形成纤维。我们研究了将K30D apoC-II与其他突变体混合是否会克服这种纤维形成能力的丧失。将K30D apoC-II与野生型、D69K或双突变体（K30D/D69K）apoC-II等摩尔混合物共同孵育，促进了K30D apoC-II掺入稳定性增加的杂合纤维中。MD模拟表明，亚基间离子对相互作用的数量增加伴随着杂合纤维稳定性的增加。这些结果证明了亚基内和亚基间电荷相互作用在稳定apoC-II淀粉样纤维中的重要作用，这一过程可能是决定蛋白质形成淀粉样纤维总体能力的关键因素。
