Protein expression and refolding – A practical guide to getting the most out of inclusion bodies.pdfVIP

Protein Expression and Purification 28 (2003) 1–8 /locate/yprep Mini review Practical considerations in refolding proteins from inclusion bodies Kouhei Tsumoto,a Daisuke Ejima,b Izumi Kumagai,a and Tsutomu Arakawac,* a Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University, Sendai, Japan b Central Research Laboratories, Ajinomoto, Inc., Kawasaki, Japan c Alliance Protein Laboratories, 3957 Corte Cancion, Thousand Oaks, CA 91360, USA Received 4 September 2002, and in revised form 2 October 2002 Abstract Refolding of proteins from inclusion bodies is affected by several factors, including solubilization of inclusion bodies by dena- turants, removal of the denaturant, and assistance of refolding by small molecule additives. We will review key parameters asso- ciated with (1) conformation of the protein solubilized from inclusion bodies, (2) change in conformation and flexibility or solubility of proteins during refolding upon reduction of denaturant concentration, and (3) the effect of small molecule additives on refolding and aggregation of the proteins. 2002 Elsevier Science (USA). All rights reserved. There is a strong demand, due to expansion of ge- bilization efficiency, in the structure of the proteins in nomic sequence database, on a rapid, large-scale pro- denatured state, and in subsequent refolding. duction of recombinant proteins. The proteins thus Refolding is initiated by reducing concentration of produced are used to identify their biological functions,