Elucidation of Short Linear Motif-Based Interactions of the FERM Domains of Ezrin, Radixin, Moesin, and Merlin

The ERM (ezrin, radixin,
and moesin) family of proteins and the
related protein merlin participate in scaffolding and signaling events
at the cell cortex. The proteins share an N-terminal FERM [band four-point-one
(4.1) ERM] domain composed of three subdomains (F1, F2, and F3) with
binding sites for short linear peptide motifs. By screening the FERM
domains of the ERMs and merlin against a phage library that displays
peptides representing the intrinsically disordered regions of the
human proteome, we identified a large number of novel ligands. We
determined the affinities for the ERM and merlin FERM domains interacting
with 18 peptides and validated interactions with full-length proteins
through pull-down experiments. The majority of the peptides contained
an apparent Yx[FILV] motif; others show alternative motifs. We defined
distinct binding sites for two types of similar but distinct binding
motifs (YxV and FYDF) using a combination of Rosetta FlexPepDock computational
peptide docking protocols and mutational analysis. We provide a detailed
molecular understanding of how the two types of peptides with distinct
motifs bind to different sites on the moesin FERM phosphotyrosine
binding-like subdomain and uncover interdependencies between the different
types of ligands. The study expands the motif-based interactomes of
the ERMs and merlin and suggests that the FERM domain acts as a switchable
interaction hub.

埃兹蛋白（ezrin）、根蛋白（radixin）与膜突蛋白（moesin）组成的ERM家族蛋白，以及相关蛋白默林（merlin），参与细胞皮层的支架构建与信号转导事件。该家族蛋白均拥有一个N端的FERM（band four-point-one，即4.1-ERM）结构域，该结构域由F1、F2、F3三个亚结构域构成，带有短线性肽基序的结合位点。我们通过将ERM家族与默林的FERM结构域，针对展示人类蛋白质组内在无序区域肽段的噬菌体展示文库进行筛选，鉴定出大量新型配体。我们测定了ERM与默林的FERM结构域分别与18条肽段结合的亲和力，并通过下拉实验验证了其与全长蛋白的相互作用。多数肽段带有特征性的Yx[FILV]基序，其余肽段则带有其他替代基序。我们结合Rosetta FlexPepDock计算肽段对接流程与突变分析，明确了两类相似但不同的结合基序（YxV与FYDF）各自的独特结合位点。我们详细阐释了两类带有不同基序的肽段如何结合至膜突蛋白FERM结构域的磷酸酪氨酸结合样亚结构域的不同位点，并揭示了不同类型配体间的相互依赖关系。本研究拓展了ERM家族与默林基于基序的相互作用组，并提示FERM结构域可作为可调控的相互作用枢纽。