Dataset Information

Identification of a prototypical single-stranded uracil DNA glycosylase from Listeria innocua.

ABSTRACT: A recent phylogenetic study on UDG superfamily estimated a new clade of family 3 enzymes (SMUG1-like), which shares a lower homology with canonic SMUG1 enzymes. The enzymatic properties of the newly found putative DNA glycosylase are unknown. To test the potential UDG activity and evaluate phylogenetic classification, we isolated one SMUG1-like glycosylase representative from Listeria innocua (Lin). A biochemical screening of DNA glycosylase activity in vitro indicates that Lin SMUG1-like glycosylase is a single-strand selective uracil DNA glycosylase. The UDG activity on DNA bubble structures provides clue to its physiological significance in vivo. Mutagenesis and molecular modeling analyses reveal that Lin SMUG1-like glycosylase has similar functional motifs with SMUG1 enzymes; however, it contains a distinct catalytic doublet S67-S68 in motif 1 that is not found in any families in the UDG superfamily. Experimental investigation shows that the S67M-S68N double mutant is catalytically more active than either S67M or S68N single mutant. Coupled with mutual information analysis, the results indicate a high degree of correlation in the evolution of SMUG1-like enzymes. This study underscores the functional and catalytic diversity in the evolution of enzymes in UDG superfamily.

SUBMITTER: Li J

PROVIDER: S-EPMC5568478 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature

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Identification of a prototypical single-stranded uracil DNA glycosylase from Listeria innocua.

Li Jing J Yang Ye Y Guevara Jose J Wang Liangjiang L Cao Weiguo W

DNA repair 20170708

A recent phylogenetic study on UDG superfamily estimated a new clade of family 3 enzymes (SMUG1-like), which shares a lower homology with canonic SMUG1 enzymes. The enzymatic properties of the newly found putative DNA glycosylase are unknown. To test the potential UDG activity and evaluate phylogenetic classification, we isolated one SMUG1-like glycosylase representative from Listeria innocua (Lin). A biochemical screening of DNA glycosylase activity in vitro indicates that Lin SMUG1-like glycos ...[more]

PMID: 28719838

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Project description:Two dimensional (2D) NMR and molecular dynamics simulations have been used to determine the three dimensional (3D) structure of a hairpin DNA, d-CTA-GAGGATCC-TUTT-GGATCCT (22mer; abbreviated as U2-hairpin), which has uracil at the second position from the 5' end of the tetraloop. The(1)H resonances of this hairpin have been assigned almost completely. NMR restrained molecular dynamics and energy minimization procedures have been used to describe the 3D structure of U2-hairpin. This study establishes that the stem of the hairpin adopts a right-handed B-DNA conformation, while the T(12)and T(15)nucleotides stack upon 3' and 5' ends of the stem, respectively. Further, T(14)stacks upon both T(12)and T(15). Though U(13)partially stacks upon T(14), no stacking interaction is observed between U(13)and T(12). All the individual nucleotide bases belonging to the stem and T(12)and T(15)of the loop adopt ' anti ' conformation with respect to their sugar moiety, while the U(13)and T(14)of the loop are in ' syn ' conformation. The turning phosphate in the loop is located between T(13)and T(14). This study and a concurrent NMR structural study on yet another hairpin DNA d-CTAGAGGAATAA-TTTU-GGATCCT (22mer; abbreviated as U4-hairpin), with uracil at the fourth position from the 5' end of the tetraloop throw light upon various interactions which have been reported between Escherichia coli uracil DNA glycosylase (UDG) and uracil containing DNA. The epsilon of T(12)and alpha, beta, gamma, epsilon and zeta of U(13)and gamma of T(14), which partially influence the local conformation of U(13)in U2-hairpin are all locked in ' trans ' conformation. Such stretched out backbone conformation in the vicinity of U(13)could be the reason as to why the U2-hairpin is found to be the poor substrate for its interaction with UDG compared to the other substrates in which the uracil is at first, third and fourth positions of the tetraloop from its 5' end, as reported earlier by Vinay and Varshney. This study shows that UDG actively promotes the flipping of uracil from a stacked conformation and rules out the possibility of UDG recognizing the flipped out uracil bases.

Dataset Information

Identification of a prototypical single-stranded uracil DNA glycosylase from Listeria innocua.

Publications

Identification of a prototypical single-stranded uracil DNA glycosylase from Listeria innocua.

Similar Datasets

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