A diffusion assay for detection and quantitation of methyl-esterified proteins on polyacrylamide gels

The methyl esterification of bacterial and mammalian proteins is a subject of increasing interest and effort. Such studies in intact cells typically involve the use of [methyl-3H]methionine which is taken up and incorporated into S-adenosyl-L-methionine, the methyl donor. The level of methylation, however, is much less than the incorporation of labeled methionine directly into protein. A diffusion assay which distinguishes [3H]methionine from the base-labile [3H]methyl esters is described here. The ester linkage is hydrolyzed at high pH to release [3H]methanol from the sample which diffuses into an adjacent pool of scintillation fluid. The assay is contained in a scintillation vial which can be counted directly.