Naslov
Strategies for folding of affinity tagged proteins using GroEL and osmolytes

Autori
Katayama, Hiroo ; McGill, Mitchell ; Kearns, Kearns ; Brzozowski, Marek ; Degner, Nicholas ; Harnett, Bliss ; Kornilayev, Boris ; Matković- Čalogović, Dubravka ; Holyoak, Todd ; Calvet, James P. ; Gogol, Edward P. ; Dees, John ; Fisher, Mark T.

Izvornik
Journal of structural and functional genomics (1345-711X) 10 (2009); 57-66

Vrsta, podvrsta i kategorija rada
Radovi u časopisima, članak, znanstveni

Ključne riječi
Chaperonin ; Osmolytes ; Affinity tags ; Protein folding ; Protein stability

Sažetak
Obtaining a proper fold of affinity tagged chimera proteins can be difficult. Frequently, the protein of interest aggregates after the chimeric affinity tag is cleaved off, even when the entire chimeric construct is initially soluble. If the attached protein is incorrectly folded, chaperone proteins such as GroEL bind to the misfolded construct and complicate both folding and affinity purification. Since chaperonin/osmolyte mixtures facilitate correct folding from the chaperonin, we explored the possibility that we could use this intrinsic binding reaction to advantage to refold two difficult-to-fold chimeric constructs. In one instance, we were able to recover activity from a properly folded construct after the construct was released from the chaperonin in the presence of osmolytes. As an added advantage, we have also found that this method involving chaperonins can enable researchers to decide (1) if further stabilization of the folded product is required and (2) if the protein construct in question will ever be competent to fold with osmolytes.

Izvorni jezik
Engleski

Znanstvena područja
Kemija

POVEZANOST RADA