Circumnavigating misfolding traps in the energy landscape through protein engineering: suppression of molten globule and aggregation in carbonic anhydrase
2004 (English)In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 43, no 21, 6803-6807 p.Article in journal (Refereed) Published
The native state of the enzyme human carbonic anhydrase (HCA II) has been stabilized by the introduction of a disulfide bond, the oxidized A23C/L203C mutant. This stabilized protein variant undergoes an apparent two-state unfolding process with suppression of the otherwise stable equilibrium, molten-globule intermediate, which is normally very prone to aggregation. Stopped-flow measurements also showed that lower amounts of the transiently occurring molten globule were formed during refolding. This led to a markedly lowered tendency for aggregation during equilibrium denaturing conditions and, more importantly, to significantly higher reactivation yields upon refolding of the fully denatured protein. Thus, a general strategy to circumvent aggregation during the refolding of proteins could be to stabilize the native state of a protein at the expense of partially folded intermediates, thereby shifting the unfolding behavior from a three-state process to a two-state one.
Place, publisher, year, edition, pages
2004. Vol. 43, no 21, 6803-6807 p.
Engineering and Technology
IdentifiersURN: urn:nbn:se:liu:diva-45720DOI: 10.1021/bi049709zOAI: oai:DiVA.org:liu-45720DiVA: diva2:266616