Journal
MUTATION RESEARCH-FUNDAMENTAL AND MOLECULAR MECHANISMS OF MUTAGENESIS
Volume 766, Issue -, Pages 56-65Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.mrfmmm.2014.06.002
Keywords
DNA damage; Abasic site; TATA box; CpG island; APE1; Time resolved fluorescence spectroscopy
Funding
- Department of Science and Technology (DST), Govt. of India [SERB/LS-36/2010]
- Central Scientific and Industrial Research (CSIR), Govt. of India [09/1023(0004)/2010-EMR-1]
- IIT Patna
- DST
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Sequences like the core element of TATA box and CpG island are frequently encountered in the genome and related to transcription. The fate of repair of clustered abasic sites in such sequences of genomic importance is largely unknown. This prompted us to investigate the sequence dependence of cleavage efficiency of APE1 enzyme at abasic sites within the core sequences of TATA box and CpG island using fluorescence dynamics and reaction kinetics. Simultaneous molecular dynamics study through steady state and time resolved fluorescence spectroscopy using unique ethidium bromide dye release assay confirmed an elevated amount of abasic site cleavage of the TATA box sequence as compared to the core CpG island. Reaction kinetics showed that catalytic efficiency of APEI for abasic site cleavage of core CpG island sequence was 4 times lower as compared to that of the TATA box. Higher value of Km was obtained from the core CpG island sequence than the TATA box sequence. This suggests a greater binding effect of APE1 enzyme on TATA sequence that signifies a prominent role of the sequence context of the DNA substrate. Evidently, a faster response from APE1 was obtained for clustered abasic damage repair of TATA box core sequences than CpG island consensus sequences. The neighboring bases of the abasic sites in the complementary DNA strand were found to have significant contribution in addition to the flanking bases in modulating APE1 activity. The repair refractivity of the bistranded clustered abasic sites arise from the slow processing of the second abasic site, consequently resulting in decreased overall production of potentially lethal double strand breaks. (C) 2014 Elsevier B.V. All rights reserved.
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