This title appears in the Scientific Report :
2014
Please use the identifier:
http://dx.doi.org/10.1016/j.jbiotec.2014.06.006 in citations.
Engineered aggregation inhibitor fusion for production of highly amyloidogenic human islet amyloid polypeptide
Engineered aggregation inhibitor fusion for production of highly amyloidogenic human islet amyloid polypeptide
Human islet amyloid polypeptide (IAPP) is the major component of pancreatic amyloid deposits in type 2 diabetes. The structural conversion of IAPP from a monomeric state into amyloid assemblies is the subject of intense research. Recombinant production of IAPP is, however, difficult due to its extre...
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Personal Name(s): | Mirecka, Ewa Agnieszka |
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Gremer, Lothar / Schiefer, Stephanie / Oesterhelt, Filipp / Stoldt, Matthias / Willbold, Dieter / Hoyer, Wolfgang (Corresponding Author) | |
Contributing Institute: |
Strukturbiochemie; ICS-6 |
Published in: | Journal of biotechnology, - (2014) S. S0168-1656(14)00287-9 |
Imprint: |
Amsterdam [u.a.]
Elsevier Science
2014
|
PubMed ID: |
24928165 |
DOI: |
10.1016/j.jbiotec.2014.06.006 |
Document Type: |
Journal Article |
Research Program: |
Structural Biology |
Publikationsportal JuSER |
Human islet amyloid polypeptide (IAPP) is the major component of pancreatic amyloid deposits in type 2 diabetes. The structural conversion of IAPP from a monomeric state into amyloid assemblies is the subject of intense research. Recombinant production of IAPP is, however, difficult due to its extreme aggregation propensity. Here we describe a novel strategy for expression of IAPP in Escherichia coli, based on an engineered protein tag, which sequesters IAPP monomers and prevents IAPP aggregation. The IAPP-binding protein HI18 was selected by phage display from a β-wrapin library. Fusion of HI18 to IAPP enabled the soluble expression of the construct. IAPP was cleaved from the fusion construct and purified to homogeneity with a yield of 3mg of isotopically labeled peptide per liter of culture. In the monomeric state, IAPP was largely disordered as evidenced by far-UV CD and liquid-state NMR spectroscopy but competent to form amyloid fibrils according to atomic force microscopy. These results demonstrate the ability of the engineered β-wrapin HI18 for shielding the hydrophobic sequence of IAPP during expression and purification. Fusion of aggregation-inhibiting β-wrapins is a suitable approach for the recombinant production of aggregation-prone proteins. |