Gq. Chen et E. Gouaux, OVEREXPRESSION OF A GLUTAMATE-RECEPTOR (GLUR2) LIGAND-BINDING DOMAIN IN ESCHERICHIA-COLI - APPLICATION OF A NOVEL PROTEIN-FOLDING SCREEN, Proceedings of the National Academy of Sciences of the United Statesof America, 94(25), 1997, pp. 13431-13436
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.