The multiple forms of lactate dehydrogenase that occur in trout were partially purified and examined by physical, catalytic, and immunological methods. Two major groups of isoenzymes were detected, which we refer to as the heart and muscle groups. Upon starch gel electrophoresis at pH 6, extracts prepared from white skeletal muscle gave rise to five closely spaced bands of lactate dehydrogenase activity with low mobility. Heart extracts exhibited another group of five (or, in some individuals, nine) closely spaced isoenzymes, which migrated rapidly toward the anode. Liver extracts contained predominantly one isoenzyme corresponding in mobility to one of the heart isoenzymes. When the lactate dehydrogenases of different species (brook, brown, and rainbow trout) were compared, differences were observed in the spacing between individual isoenzymes within the heart and muscle groups. In addition, the mobilities of the isoenzyme groups differed from species to species. To determine whether the trout heart and muscle groups are homologous with the heart and muscle lactate dehydrogenases of higher vertebrates, a comparison was made with crystalline preparations of the pure H4 (heart) and M4 (muscle) homotetramers obtained from the chicken. The two isoenzyme groups of trout were similar in molecular size to the chicken enzymes, as judged by gel filtration experiments with a calibrated column of Sephadex G-200. The heart group, like the chicken H4 enzyme, survived heating at 65°, whereas the chicken M4 enzyme and the trout muscle group were inactivated rapidly by heating at 55°. The trout heart group was more susceptible to inhibition by pyruvate than the muscle group, a difference which is similar in magnitude and direction to that known to exist between the chicken H4 and M4 enzymes. Finally, the trout heart and muscle groups were tested for reactivity with specific antisera prepared in rabbits. Antisera prepared against the chicken H4 enzyme reacted with the trout heart group but not with the trout muscle isoenzymes. The reactions were detected by immunodiffusion, enzyme inhibition, and micro-complement fixation tests. Similarly, antisera prepared against the chicken M4 enzyme reacted with the trout muscle lactate dehydrogenases but not with the trout heart enzymes. Thus, by physical, catalytic, and immunological criteria, the trout heart and muscle groups of lactate dehydrogenases appear to be homologous with the H4 and M4 enzymes, respectively, of higher vertebrates. In all the above tests, the trout liver isoenzyme behaved as a member of the heart group of lactate dehydrogenases, not as a member of the muscle group. The inclusion of the liver isoenzyme in the heart group was confirmed by experiments with an antiserum prepared against purified trout liver lactate dehydrogenase. These results, together with the published results of genetic experiments, appear to be consistent with the postulate that two distinct genes dictate the formation of the heart group of lactate dehydrogenases in trout.
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Apr 1, 1974
R L Curry + 3
Open Access
