High affinity binding of hydrophobic and autoantigenic regions of proinsulin to the 70 kDa chaperone DnaK
High affinity binding of hydrophobic and autoantigenic regions of proinsulin to the 70 kDa chaperone DnaK
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DOI：
10.1186/1471-2091-11-44
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发表时间：
2010-11-08
期刊：
影响因子：
--
通讯作者：
Kolb, Hubert
中科院分区：
文献类型：
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作者：
Burkart, Volker;Siegenthaler, Rahel K.;Kolb, Hubert
背景：分子伴侣促进肽的正确折叠，并与细胞应激期间发生的错误折叠蛋白质结合。肽与分子伴侣的复合物诱导肽定向免疫。在这里，我们分析了（前）胰岛素原与 hsp70 家族中最有特征的分子伴侣（细菌 DnaK）的相互作用。 结果：在人前胰岛素原的一组重叠 13 聚体肽中，发现信号肽和每个胰岛素原结构域中的另一个区域（A 链和 B 链、C 肽）与 DnaK 具有高亲和力结合。其中，覆盖大部分B链区域B11-23的肽表现出最强的结合，在已知的高亲和力DnaK配体范围内，解离平衡常数(K'd)为2.2+/-0.4μM。B链区域B11-23位于两个胰岛素分子之间的界面处，在胰岛素寡聚物中不可接近。事实上，天然胰岛素寡聚体显示出非常低的 DnaK 亲和力 (K'd 67.8 +/- 20.8 mu M)，而经过修饰以防止寡聚化的胰岛素原分子显示出良好的结合亲和力 (K'd 11.3 +/- 7.8 mu M)。结论：完整胰岛素仅与 hsp70 分子伴侣 DnaK 微弱相互作用，而单体胰岛素原和来自 3 个不同胰岛素原区域的肽显示出大量的分子伴侣结合。 B 链肽 B 11-23 的结合最强。有趣的是，肽 B11-23 代表 1 型糖尿病中的主要自身抗原。
Background: Chaperones facilitate proper folding of peptides and bind to misfolded proteins as occurring during periods of cell stress. Complexes of peptides with chaperones induce peptide-directed immunity. Here we analyzed the interaction of (pre)proinsulin with the best characterized chaperone of the hsp70 family, bacterial DnaK.Results: Of a set of overlapping 13-mer peptides of human preproinsulin high affinity binding to DnaK was found for the signal peptide and one further region in each proinsulin domain (A-and B-chain, C-peptide). Among the latter, peptides covering most of the B-chain region B11-23 exhibited strongest binding, which was in the range of known high-affinity DnaK ligands, dissociation equilibrium constant (K'd) of 2.2 +/- 0.4 mu M. The B-chain region B11-23 is located at the interface between two insulin molecules and not accessible in insulin oligomers. Indeed, native insulin oligomers showed very low DnaK affinity (K'd 67.8 +/- 20.8 mu M) whereas a proinsulin molecule modified to prevent oligomerization showed good binding affinity (K'd 11.3 +/- 7.8 mu M).Conclusions: Intact insulin only weakly interacts with the hsp70 chaperone DnaK whereas monomeric proinsulin and peptides from 3 distinct proinsulin regions show substantial chaperone binding. Strongest binding was seen for the B-chain peptide B 11-23. Interestingly, peptide B11-23 represents a dominant autoantigen in type 1 diabetes.