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Expressed protein ligation-mediated template protein extension

► Apolipoprotein structure and function. ► Expressed protein ligation template extension. ► Apolipoprotein A-I Milano variant as a therapeutic. Expressed protein ligation (EPL) was performed to investigate sequence requirements for a variant human apolipoprotein A-I (apoA-I) to adopt a folded struct...

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Bibliographic Details
Published in:Protein expression and purification 2012-06, Vol.83 (2), p.113-116
Main Authors: Kamei, Ayako, Hauser, Paul S., Beckstead, Jennifer A., Weers, Paul M.M., Ryan, Robert O.
Format: Article
Language:English
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Summary:► Apolipoprotein structure and function. ► Expressed protein ligation template extension. ► Apolipoprotein A-I Milano variant as a therapeutic. Expressed protein ligation (EPL) was performed to investigate sequence requirements for a variant human apolipoprotein A-I (apoA-I) to adopt a folded structure. A C-terminal truncated apoA-I, corresponding to residues 1–172, was expressed and isolated from Escherichia coli. Compared to full length apoA-I (243 amino acids), apoA-I(1–172) displayed less α-helix secondary structure and lower stability in solution. To determine if extension of this polypeptide would confer secondary structure content and/or stability, 20 residues were added to the C-terminus of apoA-I(1–172) by EPL, creating apoA-IMilano(1–192). The EPL product displayed biophysical properties similar to full-length apoA-IMilano. The results provide a general protein engineering strategy to modify the length of a recombinant template polypeptide using synthetic peptides as well as a convenient, cost effective way to investigate the structure/function relations in apolipoprotein fragments or domains of different size.
ISSN:1046-5928
1096-0279
DOI:10.1016/j.pep.2012.03.014