Repair of 3-methyladenine and 7-methylguanine in nuclear DNA of Chlamydomonas: Requirement for protein synthesis |
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Authors: | James M Sweet Betty Carda Gary D Small |
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Institution: | Section of Biochemistry, Division of Biochemistry, Physiology and Pharmacology, The University of South Dakota, Vermillion, SD 57069, U.S.A. |
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Abstract: | The removal of 3-methyladenine and 7-methylguanine from nuclear DNA was determined following exposure of Chlamydomonas reinhardi to methyl methanesulfonate (MMS). The amount of 3-methyladenine in DNA was determined using an extract from Micrococcus luteus that has a 3-methyladenine-DNA glycosylase. The amount of 7-methylguanine was estimated by heating the DNA for 30 min at 70° followed by alkaline hydrolysis of the resulting apurinic sites. The molecular weight of the DNA was determined using alkaline sucrose gradients. The 3-methyladenine is removed with a half-life of 2–3 h whereas the 7-methylaguanine is removed with a half-life of 10–12 h. The rate of removal of the 7-methylguanine is more than an order of magnitude faster than the estimated non-enzymatic hydrolysis rate indicating the probability of enzymatic repair. Addition of cycloheximide immediately after MMS treatment inhibits the removal of 3-methyladenine and 7-methylguanine from DNA. If cycloheximide is added 1.5 h after treatment with MMS, there is much less inhibition of the removal of 3-methyladenine. These results are interpreted to mean that MMS induces the synthesis of 1 or more proteins that are required for the repair of 3-methyladenine from Chlamydomonas DNA. |
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Keywords: | MMS methyl methanesulfonate |
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