Transaminase of branched chain amino acids. VII. Comparative studies on isozymes of ascites hepatoma and various normal tissues of rat.

Abstract

Rat tissues were found to contain three types of transaminase [EC 2.6.1.6] for branched chain amino acids (valine, leucine and isoleucine), enzymes I, II and III. Three enzymes could be separated by DEAE cellulose column chromatography. Enzyme I was present in all tissues examined, whereas enzyme III was only found in brain. Enzymes I and III have quite similar properties and can utilize all three branched chain amino acids about equally well as amino donors. Enzyme II has only been found in normal liver and only leucine acts as its substrate. The supernatant of Yoshida rat ascites hepatoma (AH 130) was found to contain enzymes I and III but not enzyme II. Enzyme Ills from the tumor and normal rat ”brain were purified. The preparation from the hepatoma behaved as a single protein on electrophoresis, ultracentrifugation and immunochemical diffusion. Its s0.5%20, w was estimated as 3.6 and its molecular weight as 42, 600. The enzyme Ills from hepatoma and rat brain were compared and found to be indistinguishable in their behavior on electrophoresis, immunochemical diffusion and activity neutralization, chromatography under various conditions and in various enzyme properties (substrate specificities, Km values, optimum pH and activation by 2-mercaptoethanol). Rat brain enzyme III was indistinguishable from the hog brain enzyme III im-munochemically. The latter was, however, slightly different from rat hepatoma enzyme IIL They did not crossreact in an immunochemical diffusion test, but their activities were neutralized by heterologous antiserum, though to a lesser extent than by homologous antiserum.