Overexpression in Pichia pastoris and Crystallization of an Elicitor Protein Secreted by the Phytopathogenic Fungus, Phytophthora cryptogea
Résumé
A synthetic gene encoding β-cryptogein, a member of the elicitin family, has been cloned into a vector for expression by the methylotrophic yeast,Pichia pastoris.Having first optimized the gene construction for secretion, we have overexpressed a modified β-cryptogein in a secreted form. A purification scheme suited to this expression system has been developed and highly pure, biologically active protein has been obtained. For structural analysis of this recombinant β-cryptogein, and new mutated forms thereof, optimal conditions for the crystallization of this protein have been determined and crystals that diffract to 2.2 Å have been obtained.