Protein Oxidation Implicated as the Primary Determinant of Bacterial Radioresistance (original) (raw)
Figure 2
Mn(II) Protects Proteins, but Not DNA, during In Vitro Irradiation
(A) DMSO-mediated DNA protection. pUC19 plasmid DNA was irradiated aerobically to the indicated doses in dH2O, 1% DMSO (HO• scavenger), or 5 mM MnCl2, followed by agarose gel electrophoresis (AGE). MnCl2 and DMSO were prepared in dH2O. IR, 60Co, aerobic at 0 °C. L, linear (2,686 base pairs); Lp, pUC19 + BamHI; M, size markers; OC, open circular; SC, supercoiled.
(B) Mn(II)-mediated protein protection. BamHI enzyme was irradiated aerobically to the indicated doses in dH2O, 1% DMSO, or 5 mM MnCl2, and then incubated with λ-phage DNA for 1 h at 37 °C, followed by AGE. Inset (white border), top gel (dH2O): BamHI irradiated anaerobically. M, size markers; U, uncut λ-DNA.
(C) Western blot immunoassay of protein-bound carbonyl groups in BamHI irradiated aerobically to the indicated doses in the presence or absence of 5 mM MnCl2 and/or 200 μM FeCl2. Approximately 220-ng BamHI were loaded per lane in the Western blot (W) and in the Coomassie-stained polyacrylamide denaturing gel (C); M, mixture of artificial IgG-binding protein standards; S, wide-range protein standards.