Abstract: It is intended to provide a method capable of introducing an arbitrary modification group into a synthetic protein and a method capable of easily controlling the syntheses of a modified protein and an unmodified protein. The present invention provides a method of cell-free protein synthesis comprising: performing cell-free protein synthesis using as a reaction solution of cell-free protein synthesis, a mixture solution of an extract derived from a eukaryotic cell and a reagent solution comprising as a substrate, a substance having a desired group to be introduced as a post-translational modification group; and thereby obtaining a protein comprising the post-translational modification group introduced therein. Preferably, the group to be introduced is an acyl group of fatty acid, and the form of the post-translational modification is lipid modification at the N-terminus of the protein.

Abstract: It is intended to provide a novel method for posttranslational modification in cell-free protein synthesis and a novel method for cell-free protein synthesis with the use of such posttranslational modification reaction. A method for synthesizing a protein in a cell-free system using an extract liquid for cell-free protein synthesis containing an arthropod-derived extract, characterized in that translation reaction is carried out in the presence of mammal-derived microsomal membranes.

Abstract: It is intended to provide a novel method for posttranslational modification in cell-free protein synthesis and a novel method for cell-free protein synthesis with the use of such posttranslational modification reaction. A method for synthesizing a protein in a cell-free system using an extract liquid for cell-free protein synthesis, characterized in that translation reaction is carried out in the presence of arthropod-derived microsomal membranes.

Abstract: A tumor necrosis factor (TNF) preparation with high cytolytic activity is described. The TNF preparation includes modified forms of TNF associated with or encapsulated within liposomes. The TNF molecule is modified at up to 3 amino acid residues per trimer with nearly complete retention (80-95%) of biological activity. Amino acid residues of the TNF are modified to include long chain fatty acids via TNF lysyl side chains and/or N-terminal amino acid groups. The disclosed modified TNF molecules provide a highly efficient method for preparing liposome-associated or encapsulated TNF complexes in either standard multilamellar vesicles (MLVs) or small unilamellar vesicles (SUVs) having enhanced in vivo stability. The liposomes of the present invention feature particularly small diameters in the range of 0.02-0.05 um in diameter. The binding of the modified TNF molecules to the surface of SUVs is up to 100% efficiency.