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Base substitution mutations induced in the cI gene of lambda phage by neocarzinostatin chromophore: correlation with
Abstract:
Treatment of intact lambda phage with the nonprotein chromophore of neocarzinostatin resulted in efficient phage inactivation and generation of clear-plaque mutants. Both effects required a preincubation at low pH to allow diffusion of chromophore into the phage head. Chromophore activation was then effected by addition of a sulfhydryl cofactor, followed by a shift to neutral pH. Sequence analysis of mutations mapped to the DNA-binding region of the cI gene revealed that nearly all were single base substitutions. Significant numbers of all possible base changes were found, with A:T to G:C transitions being the most frequent events. Of 11 G:C to A:T transitions, 7 were found at C residues in the trinucleotide sequence AGC, which has previously been shown to be a hotspot for chromophore-induced depyrimidination. This result, as well as the SOS dependence of mutagenesis and the overall distribution of various types of base substitutions, is consistent with the hypothesis that apurinic/apyrimidinic sites are important mutagenic lesions.
Insights
Neocarzinostatin chromophore inactivates lambda phage and causes mutations by damaging DNA. This process, dependent on low pH and sulfhydryl cofactors, creates apurinic/apyrimidinic sites, leading to base substitutions.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- Neocarzinostatin is an antibiotic with a nonprotein chromophore.
- Phage inactivation and mutagenesis are key areas in molecular biology research.
Purpose of the Study:
- To investigate the effects of neocarzinostatin chromophore on lambda phage.
- To understand the mechanism of chromophore-induced mutagenesis.
Main Methods:
- Treatment of lambda phage with neocarzinostatin chromophore under specific pH conditions.
- Sequence analysis of mutations in the phage cI gene.
- Investigating SOS dependence of mutagenesis.
Main Results:
- Neocarzinostatin chromophore efficiently inactivates lambda phage and generates clear-plaque mutants.
- Mutagenesis primarily involves single base substitutions, with a hotspot at AGC trinucleotides.
- Apurinic/apyrimidinic sites are implicated as mutagenic lesions.
Conclusions:
- The study elucidates the mechanism of neocarzinostatin chromophore-induced phage inactivation and mutagenesis.
- Apurinic/apyrimidinic site formation is a critical step in this mutagenic process.
- Findings contribute to understanding DNA damage and repair mechanisms.