Z(basic)--a novel purification tag for efficient protein recovery.
A positively charged protein domain, Z(basic), can be used as a general purification tag to achieve efficient recovery of recombinantly produced target proteins using cation-exchange chromatography. To construct a protein domain usable for ion-exchange chromatography, the surface of protein Z was engineered to be highly charged, which allowed for selective capture of target proteins on a cation-exchanger at physiological pH values. Interestingly, the novel domain, denoted Z(basic), was shown to be selective also under denaturing conditions and could preferably be used for purification of proteins solubilised from inclusion bodies. Moreover, a flexible process for solid-phase refolding was developed, using Z(basic) as a reversible linker to the cation-exchanger resin. This procedure has the inherited advantage of combining purification and refolding into a single step and still enabling elution of a concentrated product in a suitable buffer. This article summarizes development and use of the Z(basic) tag in small and pilot-plant-scale downstream processing.
Hedhammar
My
M
Department of Biotechnology, Royal Institute of Technology, Albanova University Center, Stockholm, Sweden. my.hedhammar@afb.slu.se
Hober
Sophia
S
D000595:Amino Acid Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011489:Protein Denaturation; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification*; D017386:Sequence Homology, Amino Acid; D012995:Solubility
D011506:Proteins