National Institutes of Health/National Cancer Institute (NIH/NCI)
CA100504
米国
National Institutes of Health/National Cancer Institute (NIH/NCI)
GM118009
米国
引用
ジャーナル: Nucleic Acids Res / 年: 2022 タイトル: Structural analysis of the basal state of the Artemis:DNA-PKcs complex. 著者: Go Watanabe / Michael R Lieber / Dewight R Williams / 要旨: Artemis nuclease and DNA-dependent protein kinase catalytic subunit (DNA-PKcs) are key components in nonhomologous DNA end joining (NHEJ), the major repair mechanism for double-strand DNA breaks. ...Artemis nuclease and DNA-dependent protein kinase catalytic subunit (DNA-PKcs) are key components in nonhomologous DNA end joining (NHEJ), the major repair mechanism for double-strand DNA breaks. Artemis activation by DNA-PKcs resolves hairpin DNA ends formed during V(D)J recombination. Artemis deficiency disrupts development of adaptive immunity and leads to radiosensitive T- B- severe combined immunodeficiency (RS-SCID). An activated state of Artemis in complex with DNA-PK was solved by cryo-EM recently, which showed Artemis bound to the DNA. Here, we report that the pre-activated form (basal state) of the Artemis:DNA-PKcs complex is stable on an agarose-acrylamide gel system, and suitable for cryo-EM structural analysis. Structures show that the Artemis catalytic domain is dynamically positioned externally to DNA-PKcs prior to ABCDE autophosphorylation and show how both the catalytic and regulatory domains of Artemis interact with the N-HEAT and FAT domains of DNA-PKcs. We define a mutually exclusive binding site for Artemis and XRCC4 on DNA-PKcs and show that an XRCC4 peptide disrupts the Artemis:DNA-PKcs complex. All of the findings are useful in explaining how a hypomorphic L3062R missense mutation of DNA-PKcs could lead to insufficient Artemis activation, hence RS-SCID. Our results provide various target site candidates to design disruptors for Artemis:DNA-PKcs complex formation.
名称: Protein artemis / タイプ: protein_or_peptide / ID: 2 詳細: Please use a 'blurred' map of the EMD-26192 entry to see the density of the Artemis catalytic region コピー数: 1 / 光学異性体: LEVO EC番号: 加水分解酵素; エステル加水分解酵素
凍結剤: ETHANE / チャンバー内湿度: 73 % / チャンバー内温度: 295 K 詳細: Plunge-freeze was performed using a home-made manual plunger at typical indoor humidity (Los Angeles, CA) and at room temperature..