Inhibition of initiation of protein synthesis in mammalian tissue culture cells by L-1-tosylamido-2-phenylethyl chloromethyl ketone

Abstract

Incorporation of amino acids into proteins in HeLa cells, virus-transformed 3T3 mouse fibroblasts, and mouse plasmacytoma cells is inhibited after the addition of L-1-tosylamido-2-phenylethyl chloromethyl ketone, an alkylating agent and chymotrypsin-specific protease inhibitor. Addition of this drug to tissue culture cells at concentrations of 20 to 30 mug per ml results in an irreversible inhibition of the incorporation of amino acids into cellular proteins, and a rapid and complete breakdown of polyribosomes. A comparative study examining the effects of L-1-tosylamido-2-phenylethyl chloromethyl ketone and several known inhibitors of in vivo protein synthesis, with known mechanisms of action, revealed that an optimal concentration of L-1-tosylamido-2-phenylethyl chloromethyl ketone: (a) immediately and selectively inhibits initiation of protein synthesis, (b) does not significantly affect normal elongation rates, and (c) does not promote a premature release of nascent peptides. L-1-Tosylamido-2-phenylethyl chloromenthyl ketone may prove to be a useful tool in investigating the initiatior of protein synthesis in eukaryotic cells.