Sarcomeric proteins, including myofilament and Z-disk proteins, play critical roles in regulating muscle contractile properties. A variety of isoforms and post-translational modifications (PTMs) of sarcomeric proteins have been shown to be associated with modulation of muscle functions and the occurrence of muscle diseases. Non-human primates (NHPs) are excellent research models for sarcopenia, a disease associated with alterations in sarcomeric proteins, due to their marked similarities to humans. However, the sarcomeric proteins in NHP skeletal muscle have not been well characterized. To gain a deeper understanding of sarcomeric proteins in NHP skeletal muscle, we employed top-down mass spectrometry (MS) to conduct a comprehensive analysis on isoforms and PTMs of sarcomeric proteins in rhesus macaque skeletal muscle. We identified 23 protein isoforms with 46 proteoforms of sarcomeric proteins, including 6 isoforms with 18 proteoforms from fast skeletal troponin T. Particularly, for the first time, a novel PDZ/LIM domain protein isoform, PDLIM7, was characterized with a newly identified protein sequence. Moreover, we also identified multiple PTMs on these proteins, including deamidation, methylation, acetylation, tri-methylation, phosphorylation, and S-glutathionylation. Most PTM sites were localized, including Asn13 deamidation on MLC-2S; His73 methylation on αactin; N-terminal acetylation on most identified proteins; N-terminal tri-methylation on MLC-1S, MLC-1F, MLC-2S, and MLC-2F; Ser14 phosphorylation on MLC-2S; and Ser15 and Ser16 phosphorylation on MLC-2F. In summary, a comprehensive characterization of sarcomeric proteins including multiple isoforms and PTMs in NHP skeletal muscle was achieved by analyzing intact proteins in the top-down MS approach.

中文翻译：

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非人类灵长类动物骨骼肌肌节蛋白翻译后修饰的自上而下质谱分析。

肌节蛋白，包括肌丝蛋白和 Z 盘蛋白，在调节肌肉收缩特性中起关键作用。已显示肌节蛋白的多种异构体和翻译后修饰 (PTM) 与肌肉功能的调节和肌肉疾病的发生有关。非人类灵长类动物 (NHP) 是肌肉减少症的优秀研究模型，肌肉减少症是一种与肌节蛋白改变相关的疾病，因为它们与人类有明显的相似之处。然而，NHP 骨骼肌中的肌节蛋白尚未得到很好的表征。为了更深入地了解 NHP 骨骼肌中的肌节蛋白，我们采用自上而下的质谱 (MS) 对恒河猴骨骼肌中肌节蛋白的异构体和 PTM 进行了全面分析。我们鉴定了 23 种蛋白质亚型和 46 种肌节蛋白的蛋白质型，包括来自快速骨骼肌肌钙蛋白 T 的 6 种亚型和 18 种蛋白质型。特别是，首次用新鉴定的蛋白质序列表征了一种新的 PDZ/LIM 结构域蛋白亚型 PDLIM7 . 此外，我们还鉴定了这些蛋白质上的多个 PTM，包括脱酰胺、甲基化、乙酰化、三甲基化、磷酸化和 S-谷胱甘肽化。大多数 PTM 位点是本地化的，包括 MLC-2S 上的 Asn13 脱酰胺；αactin 上的 His73 甲基化；大多数已鉴定蛋白质的 N 端乙酰化；MLC-1S、MLC-1F、MLC-2S 和 MLC-2F 上的 N 端三甲基化；MLC-2S 上的 Ser14 磷酸化；MLC-2F 上的 Ser15 和 Ser16 磷酸化。总之，