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• W3035532227 abstract "ABSTRACT Inosine is ubiquitous and essential in many biological processes, including RNA-editing. In addition, oxidative stress on RNA has been a topic of increasing interest due, in part, to its potential role in the development/progression of disease. In this work we probed the ability of three reverse transcriptases to catalyze the synthesis of cDNA in the presence of RNA templates containing inosine (I), 8-oxo-7,8-dihydroinosine (8oxo-I), guanosine (G), or 8-oxo-7,8-dihydroguanosine (8-oxoG), and explored the impact that these purine derivatives have as a function of position. To this end, we used 29-mers of RNA (as template) containing the modifications at position-18 and reverse transcribed DNA using 17-mers, 18-mers, or 19-mers (as primers). Generally reactivity of the viral RTs, MMLV / AMV / HIV, towards cDNA synthesis was similar for templates containing G or I, as well as for those with 8-oxoG or 8-oxoI. Notable differences are 1) that templates containing I enabled the incorporation of dT when using 17-mers (for exploring incorporation of dNTPs opposite the site of interest); 2) that the use of 18-mers of DNA (to explore cDNA synthesis past the lesion) led to DNA elongation inhibition in the case when a G:dA wobble pair was present, while the presence of I, 8-oxoI, or 8-oxoG led to full synthesis of the corresponding cDNA, with the latter two displaying a more efficient process; 3) that HIV-RT is more sensitive to modified base pairs in the vicinity of cDNA synthesis; and 4) that the presence of a modification two positions away from transcription initiation has an adverse impact on the overall process. Steady-state kinetics were established to determine substrate specificities towards canonical dNTPs (N = G, C, T, A). Overall we found evidence that RNA templates containing inosine are likely to incorporate dC > dT > > dA, where reactivity in the presence of dA was found to be pH dependent (process abolished at pH 7.3); and that the absence of the C2-exocyclic amine, as displayed with templates containing 8-oxoI, leads to increased selectivity towards incorporation of dA over dC. The data will be useful in assessing the impact that the presence of inosine and/or oxidatively generated lesions have on viral processes and adds to previous reports where I codes exclusively like G." @default.
• W3035532227 created "2020-06-19" @default.
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• W3035532227 date "2020-06-10" @default.
• W3035532227 modified "2023-09-26" @default.
• W3035532227 title "Translesion Synthesis by MmLV-, AMV-, and HIV-Reverse Transcriptases Using RNA Templates Containing Inosine, Guanosine, and Their 8-oxo-7,8-Dihydropurine Derivatives" @default.
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• W3035532227 doi "https://doi.org/10.1101/2020.06.10.144048" @default.
• W3035532227 hasPublicationYear "2020" @default.
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