Studies in this laboratory have demonstrated a role for methylation in chemotaxis and other important biological responses. Protein synthesis is also required for normal, sustained chemotaxis by the RAW264 macrophage cell line. One of the proteins involved in chemotactic signal transduction has been identified as Gi-2 based upon the inhibition of chemotaxis by pertussis toxin and upon the distribution of Gi-2. Gi-2 is the major pertussis toxin substrate in RAW-264 cells and is present in another chemotactic cell, the human neutrophil. We have now found that the gamma-subunit of Gi is carboxymethylated, a finding that clearly implies a regulatory role for methylation in signal transduction. Carboxymethylation occurs on the terminal alpha carboxyl group of the gamma-subunit and is accompanied by a lipidation. The carboxymethylation of a class of GTP-binding proteins distinct from the G. or Gi families has also been found. The methylated proteins have molecular weights between 20,000 and 23,000 and are found in the in membranes of RAW264 cells, rat brain, rat liver, and human buffy-coat cells. It is probable that these proteins play an important role in the regulation of specific cellular receptor systems. The methylation reactions also provide a method to identify and quantitate the methylated proteins. Both a methyl transferase and a membrane substrate from rabbit brain have been partially purified.