Potential mechanisms of photodynamic inactivation of virus by methylene blue. I. RNA-protein crosslinks and other oxidative lesions in Q beta bacteriophage.
A spectrum of oxidative lesions was observed in a bacteriophage-based model system that is very sensitive to the photodynamic activity of selected dyes. When suspensions of the intact bacteriophage Q beta were exposed to methylene blue plus light (MB + L), inactivating events, or "hits" occurred that were oxygen-dependent and that were associated with the formation of several specific lesions: (1) carbonyl moieties on proteins, (2) 8-oxo-7,8-dihydroguanine (8-oxoGua), and (3) single-strand breaks (ssb) in the RNA genome and (4) RNA-protein crosslinks. Formation of carbonyl groups associated with protein in the Q beta phage preparation correlated positively with photoinactivation of the phage with increasing doses of either of the sensitizers MB or rose bengal. Strand breaks in the Q beta genomic RNA were observable at high MB concentrations but appeared not to be significant at the lower concentrations of MB, as full-length Q beta RNA was observable well beyond the 99% inactivation point in MB dosage. It was shown that the number of 8-oxoGua lesions were unlikely to be sufficient to account for the number of lethal events. Following exposure to MB + L, crosslink formation between Q beta RNA and protein was observed by virtue of the location of RNA at the interface of phenol-aqueous extractions of phage suspensions. A significant increase over background of RNA-protein complexes (including full-length Q beta RNA) was observed at the lowest concentration of MB tested (0.5 microM), which corresponded roughly to an average of 2 lethal hits per phage or approximately 13% survival compared to the zero MB control (100% survival). Due to its close correlation with Q beta inactivation and its expected lethality, RNA-protein crosslink formation may be important as an inactivating lesion in bacteriophage Q beta following MB + L exposure.
Schneider
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JE
Oklahoma Medical Research Foundation, Free Radical Biology and Aging Research Program, Oklahoma City 73104, USA. Ed-Schneider@OMRF.OUHSC.EDU
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D017908:Allolevivirus / Q000187:drug effects* / Q000528:radiation effects; D004396:Coloring Agents / Q000494:pharmacology*; D006147:Guanine / Q000031:analogs & derivatives / Q000378:metabolism; D008027:Light; D008751:Methylene Blue / Q000494:pharmacology*; D018384:Oxidative Stress; D010777:Photochemistry; D017319:Photosensitizing Agents / Q000494:pharmacology*; D012367:RNA, Viral / Q000187:drug effects* / Q000378:metabolism / Q000528:radiation effects; D014764:Viral Proteins / Q000378:metabolism*
D004396:Coloring Agents; D017319:Photosensitizing Agents; C028696:Qbeta ribonucleic acid; D012367:RNA, Viral; D014764:Viral Proteins; C024829:8-hydroxyguanine; D006147:Guanine; D008751:Methylene Blue