Biochemical And Genetic Characteristics Of Mitochondrial Superoxide Dismutase In Grass Carp, Ctenopharyngodon Idellus

Superoxide dismutase is an evolutionarily conserved enzyme with an important role in protecting organisms and cells from the damages caused by oxygen free radicals. As a molecular clock and genetic marker, superoxide dismutase has been widely used in evolutionary studies, population genetics, and identification of different strains in the same species. However, the biochemical and genetic features of the mitochondrial superoxide dismutase have not been well characterized in fishes. In order to make this molecular marker applied in fish breeding, species protecting and evolutionary research further, we investigated the isozyme forms, biochemical genetic phenotypes. And we also study the distribution of different tissues and subunit construction of Ctenopharyngodon idellus (grass carp) mitochondrial superoxide dismutase (fm-SOD) with polyacrylamide gradient gel electrophoresis in this study. Our data revealed as follows:(1) There are three isoforms of fm-SOD which were named as fm-SOD 1, fm-SOD 2 and fm-SOD 3 according to their positions from the positive pole to the negative pole. Fm-SOD 1 is the fastest, while fm-SOD 3 is the slowest. In every different fm-SOD there are two bands, which is named as major band and sub-band. The major band migrates a little more slowly, while at the same time shows the higher activity.(2) The combination of three isoforms of fm-SOD constitutes three distinct biochemical phenotypes: phenotype 1, phenotype 2 and phenotype 3. Phenotype 1 was only associated with the fastest migratory isoform fm-SOD 1 and phenotype 3 with the slowest migratory isoform fm-SOD 3, while phenotype 2 was associated with all three isoforms. In phenotype 2, whileall of the three isoforrs all exist, fm-SOD 2 shows higher activity than that of fm-SOD 1 and fm-SOD 3, but fm-SOD 1 and fm-SOD 3 seem to be the same.(3) In the six tissues, such as heart, liver, spleen, kidney, muscle and tailfin, the three isoforms and the three phenotypes all exist. Six tissues of the same individual show the same zymogram while they have different activity: the activity of liver is highest and that of other tissues seems to be the same.(4) In wild group of grass carp, all the three phenotypes were observed, whereas in mito-gynogenetic group, only phenotypes 1 and 3 were observed. The ratio of the individual numbers of the three phenotypes is 15:51:28. After X2 test for probability, we find that the ratio of the three phenotypes in the wild group was consistent with the 1:2:1 ratio of Mendelian inheritance for 2 alleles of a single locus in the autosomal chromosomes.(5) In addition, all of the three fm-SODs were sensitive to the mixture of 15% ethanol and 25% chloroform but resistant to H2O2, indicating that the fm-SOD in grass carp is Mn-SOD.These results suggested that: (i) the mitochondrial superoxide dismutase gene in grass carp resided in chromosomes instead of mitochondrial DNA; (ii) mitochondrial superoxide dismutase was encoded by a single locus; (iii) there were two variant alleles at least in grass carp and (iv) the fm-SQD is composed of two subunits.