The incorporation of labeled lysine into the proteins of guinea pig liver homogenate.

When Cr4-labeled lysine is incubated with guinea pig liver homogenate, a-aminoadipic, ar-ketoadipic, and glutaric acids are formed from the lysine (1). These transformations were established by finding the radioactivity of the Cl4 tracer in the metabolic products. The homogenate proteins coagulated by boiling at pH 5 also contained radioactivity. The counts given by the proteins corresponded to about 0.02 to 0.03 per cent of that added as lysine; the extent of lysine incorporation into the proteins was of the same order of magnitude as Melchior and Tarver (2) had found after incubating Ss5-labeled methionine and Winnick et al. (3, 4) C4-labeled glycine with rat tissue homogenates. Yet we could not satisfy ourselves that the radioactivity remaining in the proteins in our experiments, although it persisted through exhaustive extraction, did not come from traces of adsorbed radioactive lysine. Some counts were found in the protein when the homogenate was boiled prior to incubation with isotopic lysine. The practical solution to the problem, it seemed, was to find experimental conditions in which very much more of the radioactivity added as lysine would remain in the protein after thorough washing, and little or no radioactivity in the protein of the controls; then we could conclude that the lysine was incorporated into the protein molecule and not adsorbed. The question of the mode of linkage would remain open. Eventually two sets of conditions were found in which relatively large amounts of labeled lysine are incorporated into the proteins. In the one case, with the whole homogenate as the enzyme system, the optimum pH is in the neighborhood of 6.1, and calcium is required, the optimum concentration being above 0.003 M; the reaction proceeds hardly at all without the addition of calcium. In the other case the enzyme system was the centrifugate obtained by centrifuging the diluted homogenate at 25OOg; we