EXPRESSION, PURIFICATION, AND INITIAL CHARACTERIZATION OF THE RECOMBINANT STORAGE PROTEIN-PRECURSOR OF THEOBROMA-CACAO

The gene encoding the 67-kDa cocoa storage protein precursor has been cloned from Theobroma cacao and expressed in Escherichia coli using th e pET expression system. The recombinant storage protein has been rena tured from inclusion bodies at 30 degrees C using 20 mM glycine-NaOH b uffer, pH 10.0, containing 1 mM oxidized glutathione and 0.1% Brij. Th e renatured protein was purified and demonstrated to adopt a stable na tive conformation by optical spectroscopy. Secondary structure analysi s from circular dichroism indicated the protein to be 23% alpha-helix and 38% beta-sheet, in close agreement with values obtained using a se condary structure prediction program. (C) 1996 Academic Press, Inc.