Abstract
Tyrosyl-DNA phosphodiesterase 1 (Tdp1) catalyzes the hydrolysis of the phosphodiester linkage between the DNA 3′ phosphate and a tyrosine residue as well as a variety of other DNA 3′ damaged termini. Recently we have shown that Tdp1 can liberate the 3′ DNA phosphate termini from apurinic/apyrimidinic (AP) sites. Here, we found that Tdp1 is more active in the cleavage of the AP sites inside bubble-DNA structure in comparison to ssDNA containing AP site. Furthermore, Tdp1 hydrolyzes AP sites opposite to bulky fluorescein adduct faster than AP sites located in dsDNA. Whilst the Tdp1 H493R (SCAN1) and H263A mutants retain the ability to bind an AP site-containing DNA, both mutants do not reveal endonuclease activity, further suggesting the specificity of the AP cleavage activity. We suggest that this Tdp1 activity can contribute to the repair of AP sites particularly in DNA structures containing ssDNA region or AP sites in the context of clustered DNA lesions.
Highlights
Tyrosyl-DNA phosphodiesterase 1 (Tdp1) is an enzyme that catalyzes the hydrolysis of 30-phosphotyrosyl bonds
We reported that human tyrosyl-DNA phosphodiesterase 1 (Tdp1) catalyzes the apurinic/apyrimidinic site (AP site) cleavage reaction to generate breaks with the 30- and 50-phosphate termini [7]
The ssDNA, dsDNA, DNA structure with bubble and AP sites located opposite the bulky lesions in dsDNA were used as substrates for Tdp1 (Fig. 1)
Summary
Tyrosyl-DNA phosphodiesterase 1 (Tdp1) is an enzyme that catalyzes the hydrolysis of 30-phosphotyrosyl bonds. It was shown earlier that Tdp shows preference to remove a tyrosine residue from the ends of single- or double-stranded DNA substrates in vitro rather than tyrosine located at a nick of duplex DNA [1e4]. It suggests involvement of the enzyme in the removal of the adducts from DNA ends rather than located at the internal sites of DNA. The reaction catalyzed by Tdp involves a covalent intermediate in which an active site histidine (His263 in human Tdp1) is linked by a phosphamide bond to the DNA 30-phosphate of the substrate [18,19]. Our data suggest a novel APE-independent pathway for processing AP sites
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