Enolase Isoenzymes in Mammalian Tissues

Abstract

The existence of isoenzymes of enolase (2-phospho-D-glycerate hydro-lyase, EC4.2.1.11) has been demonstrated electrophoretically, chromatographically and immunologically (Rider &Taylor, 1974). The three forms have been numbered on the basis of the electrophoretic migration of the rat enzymes, according to the recommendations of the IUPAC-IUB Commission on Biochemical Nomenclature (1972). In the rat, enolase 1 is the predominant isoenzyme in liver, kidney, spleen, testis, adipose tissues and erythrocytes, whereas skeletal muscle contains only enolase 3. Both these isoenzymes exist in heart together with the hybrid form, enolase 2. In human tissues, the isoenzyme distribution is essentially similar, except that small amounts of enolase 2 also occur in liver, and the relative proportion of the isoenzymes in heart is altered, the human organ having less enolase 1. Molecular-weight studies, using Sephadex G-I50 chromatography and polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate, have revealed that all three isoenzymes are dimers with a mol. wt. of around 90000. No significant difference in kinetic parameters has yet been found. Antisera raised in the chicken against the partially purified rat enolases 1 and 3 show that no immunological cross-reaction occurs between these two isoenzymes, whereas enolase 2 is susceptible to both antisera. It is concluded from these structural and immunological data that the observed heterogeneity of mammalian enolase is due to existence of two independent genetic loci, a and a. Thus enolase 1 is a dimer of two a subunits, enolase 3 is a dimer of two bsubunits, and enolase 2 is the aa dimer (Rider & Taylor, 1975). Developmental changes in the isoenzyme composition of rat heart and muscle were investigated by pooling tissues taken from litters whose ages had been determined by daily inspection of the cages. The enolase activity was determined and immunochemical titrations were carried out exactly as described previously (Rider & Taylor, 1974). The total enolase activity of foetal heart is the same as that of the adult tissue, 50 units/g wet wt., although there is a decrease to around 35 units/g between 10 and 30 days after birth. The developing muscle however, shows a continuous tenfold increase in enolase activity in the first 50 days of life from the foetal value of 20 units/g wet wt., a change similar to that previously reported by Hommes & Wilmink (1968). Both foetal heart and muscle contain enolase 1 as the predominant isoenzyme, and in both tissues the proportion of enolase 3 increases in the neonatal period. In muscle there is a rapid transition, for by the fifth day enolase 3 is the predominant form, whereas heart shows a more gradual and limited increase in the proportion of subunits, such that at 80 days of life enolase 1 still accounts for half the total tissue enolase activity. A further difference between the two tissues is that whereas the isoenzyme proportions in the heart approximate to a binomial distribution throughout development, this is not the case in the muscle. A phylogenetic study in which the muscle, heart and liver enolases from a wide range of species were tested against the two specific anti-(rat enolase) sera by the previously