HOIL-1, a component of the Linear Ubiquitin Assembly Complex (LUBAC), ubiquitylates serine and threonine residues in proteins, forming ester bonds (Kelsall et al, 2019, PNAS 116, 13293-13298). Here we report that mice expressing the E3 ligase-inactive HOIL-1[C458S] mutant accumulate polyglucosan in brain, cardiac muscle and other organs, indicating that HOIL-1’s E3 ligase activity is essential to prevent these toxic polysaccharide deposits from accumulating. We found that HOIL-1 monoubiquitylates glycogen and α1:4-linked maltoheptaose in vitro and identify the C6 hydroxyl moiety of glucose as the site of ester-linked ubiquitylation. The HOIL-1-catalysed monoubiquitylation of maltoheptaose was accelerated >100-fold by Met1-linked or Lys63-linked ubiquitin oligomers, which interact with the catalytic RBR domain of HOIL-1. HOIL-1 also transferred preformed ubiquitin oligomers to maltoheptaose en bloc, producing polyubiquitylated maltoheptaose in one catalytic step. The Sharpin and HOIP components of LUBAC, but not HOIL-1, bound to amylose resin in vitro, suggesting a potential function in targeting HOIL-1 to unbranched glucosaccharides in cells. We suggest that monoubiquitylation of unbranched glucosaccharides may initiate their removal by glycophagy to prevent precipitation as polyglucosan.

中文翻译：

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HOIL-1 催化的无支链糖糖泛素化及其通过泛素寡聚体的活化

HOIL-1 是线性泛素组装复合物 (LUBAC) 的一个组成部分，可泛素化蛋白质中的丝氨酸和苏氨酸残基，形成酯键（Kelsall 等，2019，PNAS 116, 13293-13298）。在这里，我们报告表达 E3 连接酶失活 HOIL-1[C458S] 突变体的小鼠在脑、心肌和其他器官中积累多聚葡聚糖，表明 HOIL-1 的 E3 连接酶活性对于防止这些有毒多糖沉积物积累至关重要。我们发现 HOIL-1在体外使糖原和 α1:4 连接的麦芽七糖单泛素化并确定葡萄糖的 C6 羟基部分是酯连接的泛素化位点。与 HOIL-1 的催化 RBR 结构域相互作用的 Met1 连接或 Lys63 连接的泛素寡聚体可将 HOIL-1 催化的麦芽七糖单泛素化加速 > 100 倍。HOIL-1也转移预成形泛素低聚物麦芽七糖整块，在一个催化步骤产生polyubiquitylated麦芽七糖。LUBAC 的 Sharpin 和 HOIP 成分，但不是 HOIL-1，在体外与直链淀粉树脂结合，表明 HOIL-1 具有将 HOIL-1 靶向细胞中未支化的葡萄糖的潜在功能。我们建议未支化的葡萄糖的单泛素化可以通过糖噬启动它们的去除，以防止沉淀为多聚葡聚糖。