Selective hydrolysis of damaged DNA by nuclease P1 |
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| Institution: | 1. Department of Biology, University of Copenhagen, Ole Maaloes Vej 5, DK-2200 Copenhagen N, Denmark;2. Biomedical Sciences Research Complex, University of St. Andrews, North Haugh, St. Andrews, Fife KY16 9ST, United Kingdom;1. College of Mathematics and Computer Science, Fuzhou University, Fuzhou 350108, PR China;2. School of Software and Electrical Engineering, Swinburne University of Technology, Melbourne, VIC 3122, Australia;3. School of Engineering and Technology, Central Queensland University, Townsville, QLD 4810, Australia;1. Genetics Lab, School of Health Sciences, Savitribai Phule Pune University, Ganeshkhind, Pune 411 007, India;2. Division of Biochemical Sciences, CSIR-National Chemical Laboratory, Dr. Homi Bhabha Road, Pune, Maharashtra 411 008, India;3. Vaccine Formulation and Research Centre, Gennova Biopharmaceuticals Limited, Pune, Maharashtra 411 057, India;4. Department of Biotechnology, Savitribai Phule Pune University, Ganeshkhind, Pune 411 007, India;5. Dr. D. Y. Patil Biotechnology and Bioinformatics Institute, Dr. D.Y. Patil Vidyapeeth, Tathawade, Pune, Maharshtra 411 033, India |
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| Abstract: | The turnover rates for hydrolysis by nuclease P1 of the 16 unmodified dideoxynucleoside monophosphates were measured. In addition, the turnover rates were measured in a variety of dideoxynucleoside monophosphates containing free radical-induced base modifications. The modified bases included cis-5,6-dihydroxy-5,6-dihydrothymine (thymine glycol), 5,6-dihydrothymine, 5-hydroxymethyuracil, 8-hydroxyguanine, 5-hydroxy-5-methylhydantoin and the formamido remnant which can be derived from either a thymine or a cytosine base. The turnover rate for dinucleoside monophosphates containing 4,8-dihydro-4-hydroxy-8-oxo-guanine modifications, which are induced by singlet oxygen, were also measured. A model was devised for the hydrolysis of DNA by nuclease P1 which uses the observed turnover rates as parameters. The model predicts the abundance of monomers and dimers as hydrolysis proceeds. Whereas the level of monomers increases monotonically, the level of each dimer first increases and then falls off. There are advantages to phosphorylating dimers, as compared with monomers, using polynucleotide kinase. Consequently this model may be of interest in connection with 32P-postlabeling applied to the measurement of DNA damage in nuclease P1 partial hydrolysates of DNA. |
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