Santos Joana A, Timinskas Kęstutis, Ramudzuli Atondaho A, Lamers Meindert H, Venclovas Česlovas, Warner Digby F, Gessner Sophia J
Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, The Netherlands.
Institute of Biotechnology, Life Sciences Center, Vilnius University, Vilnius, Lithuania.
J Biol Chem. 2025 Feb;301(2):108108. doi: 10.1016/j.jbc.2024.108108. Epub 2024 Dec 18.
The mycobacterial mutasome-comprising ImuA', ImuB, and DnaE2-has been implicated in DNA damage-induced mutagenesis in Mycobacterium tuberculosis. ImuB, which is predicted to enable mutasome function via its interaction with the β clamp, is a catalytically inactive Y-family DNA polymerase. Like some other members of the Y-family, ImuB features a recently identified amino acid motif with homology to the RecA N terminus (RecA-NT). Given the role of RecA-NT in RecA oligomerization, we hypothesized that ImuB RecA-NT mediates the interaction with ImuA', an RecA homolog of unknown function. Here, we constructed a panel of imuB alleles in which the RecA-NT was removed or mutated. Our results indicate that RecA-NT is critical for the interaction of ImuB with ImuA'. A region downstream of RecA-NT, ImuB-C, appears to stabilize the ImuB-ImuA' interaction, but its removal does not prevent complex formation. In contrast, replacing two hydrophobic residues of RecA-NT, L378 and V383, disrupts the ImuA'-ImuB interaction. To our knowledge, this is the first experimental evidence suggesting a role for RecA-NT in mediating the interaction between a Y-family member and an RecA homolog.
由ImuA'、ImuB和DnaE2组成的分枝杆菌突变体与结核分枝杆菌中DNA损伤诱导的诱变有关。ImuB预计通过其与β夹子的相互作用来实现突变体功能，它是一种催化无活性的Y家族DNA聚合酶。与Y家族的其他一些成员一样，ImuB具有一个最近鉴定出的与RecA N末端（RecA-NT）具有同源性的氨基酸基序。鉴于RecA-NT在RecA寡聚化中的作用，我们推测ImuB RecA-NT介导与功能未知的RecA同源物ImuA'的相互作用。在这里，我们构建了一组去除或突变了RecA-NT的imuB等位基因。我们的结果表明，RecA-NT对ImuB与ImuA'的相互作用至关重要。RecA-NT下游的一个区域，即ImuB-C，似乎稳定了ImuB-ImuA'的相互作用，但其去除并不妨碍复合物的形成。相反，替换RecA-NT的两个疏水残基L378和V383会破坏ImuA'-ImuB的相互作用。据我们所知，这是第一个表明RecA-NT在介导Y家族成员与RecA同源物之间相互作用中起作用的实验证据。
