Inactivation of bacteriophage phi X174 by mitomycin C in the presence of sodium hydrosulfite and cupric ions.
Bacteriophage phi X174 was inactivated by mitomycin C reduced with sodium hydrosulfite in the presence of cupric ions (Cu2+). 99% of the phage particles lost their plaque-forming abilities when incubated with 1.5 . 10(-4) M mitomycin C, 5.7 . 10(-4) M sodium hydrosulfite and 1.0 . 10(-4) M CuCl2 for 120 min at 37 degrees C in 0.05 M Tris--HCl buffer (pH 8.1). Sodium borohydride and thiol-reducing agents such as L-cysteine, 2-mercaptoethanol or dithiothreitol could not serve as a substitute for sodium hydrosulfite and other transition metal ions such as Fe2+, Fe3+, Mn2+, Co2+ and Zn2+ were of no effect. Inactivated phage sedimented at 114S just as intact phage, but phage DNA was degraded. Strand-scission was observed when phi X174 single-stranded DNA was directly reacted with mitomycin C reduced with sodium hydrosulfite in the presence of CuCl2. Phage inactivation was inhibited bycatalase, EDTA and several scavengers such as cysteamine, 2-aminoethylisothiuronium bromide HBr (AET), 4,5-dihydroxy-1,3-benzene-disulfonic acid (Tiron), or 1,4-diazabicyclo[2,2,2]octane (DABCO). These results suggest that free oxygen radicals and mitomycin C semiquinone radical generated during autoxidation of reduced mitomycin C in the presence of cupric ions cause the degradation of phy X174 DNA.
D010584:Bacteriophage phi X 174 / Q000187:drug effects* / Q000378:metabolism; D002374:Catalase / Q000494:pharmacology; D003300:Copper / Q000494:pharmacology*; D004277:DNA, Single-Stranded / Q000378:metabolism; D004279:DNA, Viral / Q000378:metabolism; D004227:Dithionite / Q000494:pharmacology*; D004492:Edetic Acid / Q000494:pharmacology; D005609:Free Radicals; D006863:Hydrogen-Ion Concentration; D008937:Mitomycins / Q000494:pharmacology*; D010100:Oxygen; D013447:Sulfites / Q000494:pharmacology*
D004277:DNA, Single-Stranded; D004279:DNA, Viral; D005609:Free Radicals; D008937:Mitomycins; D013447:Sulfites; D004227:Dithionite; D003300:Copper; D004492:Edetic Acid; D002374:Catalase; D010100:Oxygen