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. 2024 Aug 7;15:6517. doi: 10.1038/s41467-024-50836-6

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© The Author(s) 2024

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 4 — A In vitro kinase assays were performed with different concentrations of His-tagged human APE1 (His-hAPE1) as indicated, Flag-tagged WT human ATM (Flag-hATM) and His-tagged kinase-deficient human Chk2 (His-hChk2), with DMSO or ATM inhibitor KU55933 (1 mM). ATM kinase activity was indicated by Chk2 phosphorylation (P-hChk2) via immunoblotting using anti-Chk2 T68 phosphorylation specific antibodies. B Kinase assays were performed with different concentrations of His-hAPE1 as indicated, WT or KD (kinase-dead) Flag-hATM and His-hChk2. ATM kinase activity was indicated by P-hChk2. C Kinase assays were performed with Control (PBS), GST, WT or ΔNT34 GST-Xenopus APE1, Flag-hATM and His-hChk2, with the addition of DMSO or KU55933 (1 mM). ATM kinase activity was indicated by P-hChk2. D Kinase assays were performed with Control (PBS), GST, GST-Xenopus APE1 WT or ΔNT34, Flag-tagged WT or kinase-dead (KD) ATM and His-hChk2. ATM kinase activity was indicated by P-hChk2. E Quantification and statistical analysis of the ratio of P-hChk2 vs. hChk2 from Panel C. Data are presented as mean values ± SD. **p(GST vs GST-APE1-WT) = 0.0001; **p(GST-APE1-WT vs GST-APE1-ΔNT34) = 0.0016; two-tailed, unpaired t-test, n = 3. F Kinase assays were performed with different concentrations of His-tagged APE1 NT34 (His-APE1-NT34) and DMSO/KU55933 as indicated. Chk2 phosphorylation was examined via immunoblotting analysis. The data presented in Panel A–D and F are representative of three biological replicates. Source data are provided as a Source Data file.