Santamaria Andreas, Pereira Daniel, Pawar Nisha, Kelly Bernard T, Carrascosa-Tejedor Javier, Sardo Mariana, Mafra Luís, Fragneto Giovanna, Owen David J, Marín-Montesinos Ildefonso, Guzmán Eduardo, Zaccai Nathan R, Maestro Armando
Institut Laue-Langevin, 71 Avenue des Martyrs, 38042, Grenoble, France.
Departamento de Química-Física, Facultad de Ciencias Químicas, Universidad Complutense, Ciudad Universitaria s/n, Madrid, Spain.
Commun Chem. 2025 Jul 29;8(1):219. doi: 10.1038/s42004-025-01590-3.
Clathrin assembly lymphoid myeloid leukaemia protein (CALM) is involved in the formation of clathrin-mediated endocytic coats by virtue of binding many proteins involved in the process, including clathrin itself and AP2 cargo adaptor complex. CALM is able to specifically recognize the inner leaflet of the plasma membrane by binding the membrane's phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P). Here, a quantitative biophysical approach -combining neutron/X-ray scattering, solid-state NMR, atomic force microscopy, and quartz crystal microbalance with dissipation monitoring-, was exploited to investigate CALM interaction with PtdIns(4,5)P-presenting model membranes. The presented experimental data reveal CALM's folded domain partially embeds (12% volume occupancy) within the membrane, directly coordinating a cluster of 4 to 5 PtdIns(4,5)P molecules via phosphate interactions. The N-terminal amphipathic helix inserts ~8 Å into the headgroup region, reducing local membrane stiffness by 36% (from 22 to 14 MPa) while increasing viscoelastic dissipation. These results establish a plausible threefold curvature-generation mechanism: PtdIns(4,5)P clustering, helix insertion-induced lipid compaction and global mechanical softening-collectively lowering the energy barrier for membrane deformation.
网格蛋白组装淋巴样髓样白血病蛋白（CALM）通过结合许多该过程中涉及的蛋白质，包括网格蛋白本身和AP2货物衔接复合体，参与网格蛋白介导的内吞衣被的形成。CALM能够通过结合细胞膜的磷脂酰肌醇4,5-二磷酸（PtdIns(4,5)P）特异性识别质膜的内小叶。在此，采用了一种定量生物物理方法——结合中子/ X射线散射、固态核磁共振、原子力显微镜和带耗散监测的石英晶体微天平——来研究CALM与呈现PtdIns(4,5)P的模型膜的相互作用。所呈现的实验数据表明，CALM的折叠结构域部分嵌入（体积占有率为12%）膜内，通过磷酸相互作用直接协调4至5个PtdIns(4,5)P分子的簇。N端两亲螺旋插入头基团区域约8 Å，使局部膜硬度降低36%（从22 MPa降至14 MPa），同时增加粘弹性耗散。这些结果建立了一种合理的三重曲率产生机制：PtdIns(4,5)P聚集、螺旋插入诱导的脂质压实和整体机械软化——共同降低膜变形的能垒。
