Improved non-chromatographic purification of a recombinant protein by cationic elastin-like polypeptides.
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.
Lim
Dong Woo
DW
Department of Biomedical Engineering, Box 90281, Duke University, Durham, North Carolina 27708-0281, USA.
Trabbic-Carlson
Kimberly
K
Mackay
J Andrew
JA
Chilkoti
Ashutosh
A
D000595:Amino Acid Sequence; D001483:Base Sequence; D002412:Cations / Q000737:chemistry; D004549:Elastin / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics; D008969:Molecular Sequence Data; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification*; D012965:Sodium Chloride / Q000737:chemistry; D013879:Thioredoxins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification*
D002412:Cations; D019151:Peptide Library; D010455:Peptides; D011993:Recombinant Fusion Proteins; D012965:Sodium Chloride; D013879:Thioredoxins; D004549:Elastin