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Site‐specific functionality and tryptophan mimicry of lipidation in tetraspanin CD9
The FEBS Journal ( IF 5.5 ) Pub Date : 2020-03-17 , DOI: 10.1111/febs.15295 Viviana Neviani 1 , Sjoerd van Deventer 2 , Tobias P. Wörner 3 , Katerina T. Xenaki 4 , Michiel van de Waterbeemd 3 , Remco N.P. Rodenburg 1 , Inge M.N. Wortel 2 , Jeroen K. Kuiper 2 , Sofie Huisman 1 , Joke Granneman 1 , Paul M.P. van Bergen en Henegouwen 4 , Albert J.R. Heck 3 , Annemiek B. van Spriel 2 , Piet Gros 1
The FEBS Journal ( IF 5.5 ) Pub Date : 2020-03-17 , DOI: 10.1111/febs.15295 Viviana Neviani 1 , Sjoerd van Deventer 2 , Tobias P. Wörner 3 , Katerina T. Xenaki 4 , Michiel van de Waterbeemd 3 , Remco N.P. Rodenburg 1 , Inge M.N. Wortel 2 , Jeroen K. Kuiper 2 , Sofie Huisman 1 , Joke Granneman 1 , Paul M.P. van Bergen en Henegouwen 4 , Albert J.R. Heck 3 , Annemiek B. van Spriel 2 , Piet Gros 1
Affiliation
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Lipidation of transmembrane proteins regulates many cellular activities, including signal transduction, cell–cell communication, and membrane trafficking. However, how lipidation at different sites in a membrane protein affects structure and function remains elusive. Here, using native mass spectrometry we determined that wild‐type human tetraspanins CD9 and CD81 exhibit nonstochastic distributions of bound acyl chains. We revealed CD9 lipidation at its three most frequent lipidated sites suffices for EWI‐F binding, while cysteine‐to‐alanine CD9 mutations markedly reduced binding of EWI‐F. EWI‐F binding by CD9 was rescued by mutating all or, albeit to a lesser extent, only the three most frequently lipidated sites into tryptophans. These mutations did not affect the nanoscale distribution of CD9 in cell membranes, as shown by super‐resolution microscopy using a CD9‐specific nanobody. Thus, these data demonstrate site‐specific, possibly conformation‐dependent, functionality of lipidation in tetraspanin CD9 and identify tryptophan mimicry as a possible biochemical approach to study site‐specific transmembrane‐protein lipidation.
中文翻译：
四跨膜蛋白CD9中脂化的位点特异性功能和色氨酸模拟
跨膜蛋白的脂质调节许多细胞活动，包括信号转导，细胞间通讯和膜运输。但是，膜蛋白不同部位的脂化如何影响结构和功能仍然不清楚。在这里，我们使用自然质谱法确定了野生型人四跨膜蛋白CD9和CD81表现出结合的酰基链的非随机分布。我们发现在其三个最频繁的脂化部位CD9脂化足以满足EWI-F的结合，而半胱氨酸-丙氨酸CD9突变则明显降低了EWI-F的结合。通过将所有三个或最常见的脂化位点突变成色氨酸，可以挽救CD9与EWI-F的结合。这些突变不会影响CD9在细胞膜中的纳米级分布，如使用CD9特异性纳米抗体的超分辨率显微镜所示。因此，这些数据证明了四跨膜蛋白CD9中脂化的位点特异性（可能是构象依赖性）功能，并将色氨酸模拟物作为研究位点特异性跨膜蛋白脂化的一种可能的生化方法。
更新日期：2020-03-17
中文翻译：
四跨膜蛋白CD9中脂化的位点特异性功能和色氨酸模拟
跨膜蛋白的脂质调节许多细胞活动，包括信号转导，细胞间通讯和膜运输。但是，膜蛋白不同部位的脂化如何影响结构和功能仍然不清楚。在这里，我们使用自然质谱法确定了野生型人四跨膜蛋白CD9和CD81表现出结合的酰基链的非随机分布。我们发现在其三个最频繁的脂化部位CD9脂化足以满足EWI-F的结合，而半胱氨酸-丙氨酸CD9突变则明显降低了EWI-F的结合。通过将所有三个或最常见的脂化位点突变成色氨酸，可以挽救CD9与EWI-F的结合。这些突变不会影响CD9在细胞膜中的纳米级分布，如使用CD9特异性纳米抗体的超分辨率显微镜所示。因此，这些数据证明了四跨膜蛋白CD9中脂化的位点特异性（可能是构象依赖性）功能，并将色氨酸模拟物作为研究位点特异性跨膜蛋白脂化的一种可能的生化方法。
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