Hub proteins participate in cellular regulation by dynamic binding of multiple proteins within interaction networks. The hub protein LC8 reversibly interacts with more than 100 partners through a flexible pocket at its dimer interface. To explore the diversity of the LC8 partner pool, we screened for LC8 binding partners using a proteomic phage display library composed of peptides from the human proteome, which had no bias toward a known LC8 motif. Of the identified hits, we validated binding of 29 peptides using isothermal titration calorimetry. Of the 29 peptides, 19 were entirely novel, and all had the canonical TQT motif anchor. A striking observation is that numerous peptides containing the TQT anchor do not bind LC8, indicating that residues outside of the anchor facilitate LC8 interactions. Using both LC8-binding and nonbinding peptides containing the motif anchor, we developed the "LC8Pred" algorithm that identifies critical residues flanking the anchor and parses random sequences to predict LC8-binding motifs with ∼78% accuracy. Our findings significantly expand the scope of the LC8 hub interactome.

中文翻译：

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系统识别毂蛋白LC8的识别基序。

集线器蛋白通过相互作用网络中多种蛋白的动态结合来参与细胞调节。枢纽蛋白LC8通过其二聚体界面的柔性口袋可逆地与100多个伴侣相互作用。为了探索LC8伴侣库的多样性，我们使用蛋白质组噬菌体展示文库筛选了LC8结合伴侣，该文库由来自人类蛋白质组的肽组成，对已知的LC8基序没有偏见。在确定的结果中，我们使用等温滴定量热法验证了29种肽的结合。在这29种肽中，有19种是全新的，并且都具有规范的TQT基序锚。令人惊讶的观察是，许多含有TQT锚的肽不结合LC8，表明锚之外的残基促进了LC8相互作用。我们同时使用包含基序锚的LC8结合肽和非结合肽，开发了“ LC8Pred”算法，该算法可识别锚定侧翼的关键残基，并解析随机序列以〜78％的准确性预测LC8结合基序。我们的发现大大扩展了LC8集线器相互作用组的范围。