Improved Non-chromatographic Purification of a Recombinant Protein by Cationic Elastin-like Polypeptides

• Published in: Biomacromolecules 2007-05, Vol.8 (5), p.1417-1424
• Main Authors: Lim, Dong Woo, Trabbic-Carlson, Kimberly, MacKay, J. Andrew, Chilkoti, Ashutosh
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Base Sequence; Biological and medical sciences; Biotechnology; Cations - chemistry; Elastin - biosynthesis; Elastin - chemistry; Elastin - genetics; Escherichia coli - chemistry; Escherichia coli - genetics; Fundamental and applied biological sciences. Psychology; Methods. Procedures. Technologies; Molecular Sequence Data; Others; Peptide Library; Peptides - chemistry; Peptides - genetics; Recombinant Fusion Proteins - biosynthesis; Recombinant Fusion Proteins - chemistry; Recombinant Fusion Proteins - isolation & purification; Sodium Chloride - chemistry; Thioredoxins - biosynthesis; Thioredoxins - chemistry; Thioredoxins - isolation & purification; Various methods and equipments
• ISSN: 1525-7797; 1526-4602
• DOI: 10.1021/bm060849t

This paper reports an improvement in the purification of thioredoxin (Trx) expressed from E. coli by inverse transition cycling (ITC) using cationic elastin-like polypeptides (ELPs). Two ELP libraries having 2% and 5% lysine residues and molecular weights ranging from 4 to 61.1 kDa showed greater salt sensitivity in their inverse transition behavior than purely aliphatic ELPs. Expression yield of Trx-ELP fusions was an unpredictable function of guest residue composition, but reducing the molecular weight of the ELP tag generally increased Trx yield. A cationic 4.3 kDa ELP is the shortest ELP used to purify any protein by ITC to date. A 15.9 kDa ELP with a guest residue composition of K:V:F of 1:7:1 was found to be the optimal cationic tag to purify Trx, as it provided 50% greater Trx yield and only required one-fifth the added NaCl for purification of Trx as compared to previously used aliphatic ELP tags.