OVEREXPRESSION OF A GLUTAMATE-RECEPTOR (GLUR2) LIGAND-BINDING DOMAIN IN ESCHERICHIA-COLI - APPLICATION OF A NOVEL PROTEIN-FOLDING SCREEN

Expression of the S1S2 ligand binding domain [Kuusinen, A., Arvola, M. & Keinanen, K. (1995) EMBO J 14, 6327-6332] of the rat lpha-amino-3-h ydroxy-5-methylisoxazole-3-propionic acid-selective glutamate receptor GluR2 in Escherichia coli under control of a T7 promoter leads to pro duction of >100 mg/liter of histidine-tagged S1S2 protein (HS1S2) in t he form of inclusion bodies. Using a novel fractional factorial foldin g screen and a rational, step-by-step approach, multiple conditions we re determined for the folding of the HS1S2 lpha-amino-3-hydroxy-5-meth ylisoxazole-4-propionic acid binding domain. Characterization of the H S1S2 ligand binding domain showed that it is water-soluble, monomeric, has significant secondary structure, and is sensitive to trypsinolysi s at sites close to the beginning of the putative transmembrane region s. Application of a fractional factorial folding screen to other prote ins may provide a useful means to evaluate E. coli as an economical an d convenient expression host.