Zbasic : A Purification Tag for Selective Ion-Exchange Recovery

• A positively charged protein domain, denoted Zbasic, can be used as a general purification tag for purification of recombinantly produced target proteins by cation-exchange chromatography. The Zbasic domain is constructed from the Protein A-derived Z-domain, and engineered to be highly charged, which allows selective capture on a cation exchanger at physiological pH values. Moreover, Zbasic is selective also under denaturing conditions and can be used for purification of proteins solubilized from inclusion bodies. Zbasic can then be used as a flexible linker to the cation-exchanger resin, and thereby allows solid-phase refolding of the target protein. Herein, protocols for purification of soluble Zbasic-tagged fusion proteins, as well as for integrated purification and solid-phase refolding of insoluble fusion proteins, are described. In addition, a procedure for enzymatic tag removal and recovery of native target protein is outlined.

Ämnesord

NATURVETENSKAP  -- Biologi -- Biokemi och molekylärbiologi (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences -- Biochemistry and Molecular Biology (hsv//eng)

Nyckelord

Fusion tag

Ion-exchange chromatography

Protein A

Proteolytic cleavage

Solid-phase refolding

Zbasic

fusion protein

protein tag

proteinase

unclassified drug

z basic fusion protein

amino acid sequence

amino terminal sequence

cation exchange

cell inclusion

expanded bed adsorption

expression vector

ion exchange chromatography

nucleotide sequence

polyacrylamide gel electrophoresis

protein degradation

protein expression

protein isolation

protein purification

protein refolding

protein targeting

Publikations- och innehållstyp

ref (ämneskategori)

kap (ämneskategori)