Novel method for screening of radioprotective agents providing protection to DNA ligase against gamma radiation induced damage.

PURPOSE: A simple, sensitive and novel method was developed to screen out potential agents able to protect functional activity of DNA ligase against gamma irradiation-induced damage. Repeatability, authenticity and sensitivity of the method was verified by analyzing DNA ligase protecting activities of well-known radioprotectors such as amifostine, trolox, melatonin, semiquinone glucoside derivative (SQGD) and an antioxidant gallic acid in extremely low concentration (1 µg/reaction). MATERIAL AND METHODS: Two different sets (Set A and B) of T4 DNA ligase (1 unit/set) were prepared. Set 'A' (negative control) was exposed to different doses (3-5 kGy) of gamma radiation in the absence of radioprotective compounds. Set B (test) was exposed to similar doses of gamma radiation in the presence of radioprotective compounds. Following irradiation, DNA ligase was mixed with λ DNA (250 ng) pre-digested with Hind III restriction endonuclease. Ligation reaction was performed in both sets simultaneously at 22°C for 20 min and reaction product was analyzed using agarose gel electrophoresis. RESULTS: Complete DNA ligation was observed in samples where DNA ligase was irradiated in the presence of radioprotectective compounds, i.e., amifostine, trolox, melatonin and a natural radioprotector semiquinone glucoside derivative (SQGD) individually, while, functional impairment in ligation activity of DNA ligase was evident in samples in which DNA ligase was irradiated in the absence of a radioprotective compound. CONCLUSION: The current method was able to provide significant input to screen out radioprotective compounds able to protect DNA ligase functional activity against gamma radiation-induced functional impairment.

The viral potassium channel Kcv: structural and functional features.

The chlorella virus PBCV-1 was the first virus found to encode a functional potassium channel protein (Kcv). Kcv is small (94 aa) and basically consists of the M1-P-M2 (membrane-pore-membrane) module typical of the pore regions of all known potassium channels. Kcv forms functional channels in three heterologous systems. This brief review discusses the gating, permeability and modulation properties of Kcv and compares them to the properties of bacterial and mammalian K+ channels.