The crystal structure of the RhoA-AKAP-Lbc DH-PH domain complex.
The crystal structure of the RhoA-AKAP-Lbc DH-PH domain complex.
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DOI：
10.1042/bj20140606
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发表时间：
2014-12-01
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影响因子：
--
通讯作者：
Elkins JM
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文献类型：
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作者：
Abdul Azeez KR;Knapp S;Fernandes JM;Klussmann E;Elkins JM
AKAP-Lbc（A-激酶锚定蛋白-Lbc，也称为AKAP 13）的RhoGEF（Rho GT鸟嘌呤核苷酸交换因子）结构域催化RhoA上的核苷酸交换，并参与心脏肥大的发生。AKAP-Lbc的RhoGEF活性也与癌症有关。我们已经确定了RhoA-GDP和AKAP-Lbc RhoGEF [DH（Dbl-homologous）-PH（pleckstrin homologous）]结构域之间的复合物的X射线晶体结构，分辨率为2.1 nm（1 nm =0.1 nm）。与相关的RhoGEF蛋白如白血病相关的RhoGEF相比，其结构揭示了重要的差异。比较DH-PH结构域与单独的DH结构域的活性的核苷酸交换测定显示PH结构域在核苷酸交换中没有作用，这通过RhoA-AKAP-Lbc结构来解释。与分离的AKAP-Lbc DH结构域的结构的比较揭示了N-末端“GEF开关”区域在与RhoA结合时构象的变化。等温滴定量热法显示AKAP-Lbc仅对RhoA具有微摩尔亲和力，这与AKAP-Lbc上小分子的潜在结合口袋的存在相结合，提高了用GEF抑制剂靶向AKAP-Lbc的可能性。与RhoA复合的AKAP-Lbc的RhoGEF结构域的晶体结构结合核苷酸交换测定解释了与相关RhoGEF蛋白的差异，并允许用小分子靶向AKAP-Lbc RhoGEF结构域的可能性。
The RhoGEF (Rho GTPase guanine-nucleotide-exchange factor) domain of AKAP-Lbc (A-kinase-anchoring protein-Lbc, also known as AKAP13) catalyses nucleotide exchange on RhoA and is involved in the development of cardiac hypertrophy. The RhoGEF activity of AKAP-Lbc has also been implicated in cancer. We have determined the X-ray crystal structure of the complex between RhoA–GDP and the AKAP-Lbc RhoGEF [DH (Dbl-homologous)–PH (pleckstrin homology)] domain to 2.1 Å (1 Å=0.1 nm) resolution. The structure reveals important differences compared with related RhoGEF proteins such as leukaemia-associated RhoGEF. Nucleotide-exchange assays comparing the activity of the DH–PH domain to the DH domain alone showed no role for the PH domain in nucleotide exchange, which is explained by the RhoA–AKAP-Lbc structure. Comparison with a structure of the isolated AKAP-Lbc DH domain revealed a change in conformation of the N-terminal ‘GEF switch’ region upon binding to RhoA. Isothermal titration calorimetry showed that AKAP-Lbc has only micromolar affinity for RhoA, which combined with the presence of potential binding pockets for small molecules on AKAP-Lbc, raises the possibility of targeting AKAP-Lbc with GEF inhibitors. The crystal structure of the RhoGEF domain of AKAP-Lbc in complex with RhoA combined with nucleotide exchange assays explain differences to related RhoGEF proteins and allow the possibility of targeting the AKAP-Lbc RhoGEF domain with small molecules.