TY  - JOUR
AU  - Mirecka, Ewa Agnieszka
AU  - Gremer, Lothar
AU  - Schiefer, Stephanie
AU  - Oesterhelt, Filipp
AU  - Stoldt, Matthias
AU  - Willbold, Dieter
AU  - Hoyer, Wolfgang
TI  - Engineered aggregation inhibitor fusion for production of highly amyloidogenic human islet amyloid polypeptide
JO  - Journal of biotechnology
VL  - -
SN  - 0168-1656
CY  - Amsterdam [u.a.]
PB  - Elsevier Science
M1  - FZJ-2014-05667
SP  - S0168-1656(14)00287-9
PY  - 2014
AB  - 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.
LB  - PUB:(DE-HGF)16
UR  - <Go to ISI:>//WOS:000345684000028
C6  - pmid:24928165
DO  - DOI:10.1016/j.jbiotec.2014.06.006
UR  - https://juser.fz-juelich.de/record/172157
ER  -