Fig. 5: The structure of the K63A/5’-p-ADPR-DNA complex.
From: Crystal structures and snapshots along Tpt1-catalyzed phosphate transfer from nucleic acid to NAD+
a In vitro assays showing DNA 5’-PO42- transfer activities of wild-type and mutant proteins of TkoTpt1. Experiments were repeated independently three times with similar results. Source data are provided in the Source Data file. b Time course analysis of the DNA 5’-PO42- transfer activities of TkoTpt1 K63A mutant. The reaction system contains 0.25 μM K63A mutant, 0.5 μM substrate DNA, and 2 mM NAD+ in reaction buffer. The distributions of the 5′-p-ADPR DNA intermediate and 5′-OH DNA product (expressed as percentage of total DNA) are plotted as a function of reaction time. Each datum in the graphs is the average of three independent experiments ±SEM. c Overall folding of the K63A/5’-p-ADPR-DNA complex. The key 5’-p-ADPR-DNA intermediate is shown as sticks. d 2Fo-Fc electron density maps of the intermediate bound in the K63A/5’-p-ADPR-DNA complex. e The detailed interactions between TkoTpt1 and the phosphate groups of the bound intermediate. f Superposition showing the conformational differences between the intermediate and the product. Protein residues and the intermediate in K63A/5’-p-ADPR-DNA complex are colored as in (e). For TkoTpt1/DNA/Appr>P complex-A, protein residues, DNA, and the Appr>P product are all colored in light blue.
