PHOSPHATASE-IMMUNODEPLETED CELL-FREE PROTEIN-SYNTHESIS SYSTEM

A cell-free protein synthesis system using wheat-germ extract was impr oved by a novel approach involving selective removal of endogenous pho sphatase. which reduces both the duration and the rate of translation by hydrolyzing ATP and GTP, from the translational reaction. Immunodep letion of the phosphatases by the antibodies raised against the major one of the wheat-germ phosphatase isozymes removed 20-40% of ATP-hydro lysis activity from the wheatgerm extract, and thereby prolonged the r eaction period of translation. Moreover, the condensation of the phosp hate-immunodepleted extract by polyethylene glycol (PEG) precipitation and the addition of copper ions, which was known to inhibit phosphata se and nuclease activity, increased the protein synthesis more than tw o-fold compared with the reaction using control IgG-treated condensed extract. (C) 1998 Elsevier Science B.V. All rights reserved.