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• W2101854524 endingPage "1847" @default.
• W2101854524 startingPage "1840" @default.
• W2101854524 abstract "T4 RNA ligase 2 (Rnl2) repairs 3′-OH/5′-PO 4 nicks in duplex nucleic acids in which the broken 3′-OH strand is RNA. Ligation entails three chemical steps: reaction of Rnl2 with ATP to form a covalent Rnl2–(lysyl-Nζ)–AMP intermediate (step 1); transfer of AMP to the 5′-PO 4 of the nick to form an activated AppN– intermediate (step 2); and attack by the nick 3′-OH on the AppN– strand to form a 3′–5′ phosphodiester (step 3). Here we used rapid mix-quench methods to analyze the kinetic mechanism and fidelity of single-turnover nick sealing by Rnl2–AMP. For substrates with correctly base-paired 3′-OH nick termini, k step2 was fast (9.5 to 17.9 sec −1 ) and similar in magnitude to k step3 (7.9 to 32 sec −1 ). Rnl2 fidelity was enforced mainly at the level of step 2 catalysis, whereby 3′-OH base mispairs and oxoguanine, oxoadenine, or abasic lesions opposite the nick 3′-OH elicited severe decrements in the rate of 5′-adenylylation and relatively modest slowing of the rate of phosphodiester synthesis. The exception was the noncanonical A:oxoG base pair, which Rnl2 accepted as a correctly paired end for rapid sealing. These results underscore (1) how Rnl2 requires proper positioning of the 3′-terminal ribonucleoside at the nick for optimal 5′-adenylylation and (2) the potential for nick-sealing ligases to embed mutations during the repair of oxidative damage." @default.
• W2101854524 created "2016-06-24" @default.
• W2101854524 creator A5014112082 @default.
• W2101854524 creator A5062069748 @default.
• W2101854524 date "2013-10-24" @default.
• W2101854524 modified "2023-10-18" @default.
• W2101854524 title "Kinetic mechanism of nick sealing by T4 RNA ligase 2 and effects of 3′-OH base mispairs and damaged base lesions" @default.
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• W2101854524 doi "https://doi.org/10.1261/rna.041731.113" @default.
• W2101854524 hasPubMedCentralId "https://www.ncbi.nlm.nih.gov/pmc/articles/3884662" @default.
• W2101854524 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/24158792" @default.
• W2101854524 hasPublicationYear "2013" @default.
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