Initial Study On The Superoxide Dismutase Modified By Melatonin

Lots of brilliant achievements in as well as a variety of methods of chemical modifications on Superoxide Dismutase (SOD) have been reported worldwide in the past twenty years. Compared with natural SOD, the stability of the modified enzyme against heat, acid and degradation of the pepsin increased significantly. Furthermore, after modification the immunogenicity of natural SOD decreased or even completely eliminated. Despite the advantages mentioned above, functions of the modified SOD still need further improvement.In this study, new method was applied in the modification of SOD. Melatonin (MT), which is effective in both free radical elimination and immunity enhancement in organisms, was selected to modify the natural SOD molecule in order to obtain a multi-purpose modified SOD with increased stability.It is well known that SOD can specifically catalyze the dismutation of O2"in the metabolism of free radicals, however, some much more harmful free radicals such as OH and ONOO- could not be eliminated by SOD. While both of the above free radicals could be clear up with melatonin. Theoretically, when chemically coupled with MT, the MT - SOD combination will be able to scavenge more kinds of free radicals, and the function of SOD could be extended then.In order to couple the MT to SOD, the method of the preparation of anti-melatonin antibodies was employed. First, formaldehyde, as a coupling reagent, reacts with the dissociative amido on the surface of SOD molecule; then coupled with melatonin.Since formaldehyde of certain concentration can denature SOD, 0.250g/100ml formaldehyde solution was screened out to be the coupling reagent after a series ofexperiments.By a method for testing the modification efficiency we established, it was detected that each SOD molecule can coupled with 6-8 melatonin molecules. Meanwhile, the MT - SOD combination not only retained 60% activity of natural SOD, but also exhibit the ability to eliminate ?OH, which was impossible in the natural SOD.MT - SOD combination and natural SOD was incubated at 751 for different period of time and then their activity was detected, respectively. It was found that the activity of MT - SOD combination retained 61.3% of that at the normal temperature while the activity of natural SOD retained only 35.2%, which shows that compared with natural SOD, the stability against heat of MT - SOD combination was also increased.