Selection for nonamyloidogenic mutants of islet amyloid polypeptide (IAPP) identifies an extended region for amyloidogenicity.
Selection for nonamyloidogenic mutants of islet amyloid polypeptide (IAPP) identifies an extended region for amyloidogenicity.
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DOI：
10.1021/bi100337p
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发表时间：
2010-09-14
期刊：
影响因子：
2.9
通讯作者：
Moffet, David A.
中科院分区：
文献类型：
--
作者：
Fox, Ayano;Snollaerts, Thibaut;Casanova, Camille Errecart;Calciano, Anastasia;Nogaj, Luiza A.;Moffet, David A.
37残基多肽IAPP的聚集，作为不溶性淀粉样蛋白或小寡聚物，似乎在II型糖尿病胰腺β-胰岛细胞的死亡中起直接作用。虽然已知IAPP是II型糖尿病淀粉样蛋白的主要成分，但尚未确定负责这种聚集的分子相互作用。为了识别容易聚集的区域，我们构建了一个随机生成的IAPP点突变体库。该突变体IAPP文库在大肠杆菌中作为报告蛋白增强绿色荧光蛋白（EGFP）的遗传融合体表达。由于IAPP的聚集速度很快，无论是独立的还是与EGFP融合的，融合蛋白都不能产生功能性的荧光EGFP。然而，导致非淀粉样蛋白序列的IAPP突变仍然是可溶性的，并允许EGFP折叠和荧光。通过这个筛选，我们鉴定出22个IAPP单突变，4个双突变和2个三突变，这些突变似乎比野生型人类IAPP的淀粉样变性更低。这些序列的比较表明，残基13和15-17在IAPP的主要淀粉样变区之外包含一个额外的聚集易发区域。
The aggregation of the 37-residue polypeptide IAPP, as either insoluble amyloid or as small oligomers, appears to play a direct role in the death of pancreatic β-islet cells in type II diabetes. While IAPP has been known to be the primary component of type II diabetes amyloid, the molecular interactions responsible for this aggregation have not been identified. To identify the aggregation-prone region(s), we constructed a library of randomly generated point mutants of IAPP. This mutant IAPP library was expressed in E. coli as genetic fusions to the reporter protein enhanced green fluorescent protein (EGFP). Because IAPP aggregates rapidly, both independently and when fused to EGFP, the fusion protein does not yield a functional, fluorescent EGFP. However, mutations of IAPP that result in non-amyloidogenic sequences remain soluble and allow EGFP to fold and fluoresce. Using this screen, we identified 22 single mutations, 4 double mutations and 2 triple mutations of IAPP that appear to be less amyloidogenic than wild type human IAPP. A comparison of these sequences suggests residues 13 and 15–17 comprise an additional aggregation-prone region outside of the main amyloidogenic region of IAPP.
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影响因子：
2.9
作者：
Marek, Peter;Abedini, Andisheh;Raleigh, Daniel P.
通讯作者：
Raleigh, Daniel P.
影响因子：
7.7
作者：
Ritzel, Robert A.;Meier, Juris J.;Butler, Peter C.
通讯作者：
Butler, Peter C.
影响因子：
5.6
作者：
Abedini, A;Raleigh, DP
通讯作者：
Raleigh, DP
影响因子：
2.1
作者：
Baine, Michael;Georgie, Daniel S.;Shiferraw, Elelta Z.;Nguyen, Theresa P. T.;Nogaj, Luiza A.;Moffet, David A.
通讯作者：
Moffet, David A.
影响因子：
5.4
作者：
de Groot, NS;Aviles, FX;Ventura, S
通讯作者：
Ventura, S