CELL-FREE PROTEIN SYNTHESIS OF DEUTERATED PROTEINS FOR STRUCTURAL AND FUNCTIONAL STUDIES BY NMR SPECTROSCOPY

摘要

A method for cell-free synthesis of at least partially deuterated proteins, peptides or assemblies thereof, in particular for NMR studies, is described. The method comprises at least the following steps: a) preparation of a cell extract under essentially non-deuterated conditions; b) exchanging D2O into the cell extract in a buffer exchange process to form a deuterated cell extract; c) cell-free synthesis using at least the deuterated cell extract, expressed and purified RNA polymerase, at least partially deuterated amino acids and a plasimd or a linear template DNA encoding said protein or peptide; d) isolation and purification of the protein or peptide. Furthermore an extract for use in such a cell-free synthesis and a kit for use in such a cell-free protein synthesis is described