Mass spectrometry for the identification and analysis of highly complex glycosylation of therapeutic or pathogenic proteins
Mass spectrometry for the identification and analysis of highly complex glycosylation of therapeutic or pathogenic proteins
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DOI：
10.1080/14789450.2020.1769479
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发表时间：
2020-04
影响因子：
3.4
通讯作者：
Yukako Ohyama;K. Nakajima;M. Renfrow;J. Novak;Kazuo Takahashi
中科院分区：
文献类型：
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作者：
Yukako Ohyama;K. Nakajima;M. Renfrow;J. Novak;Kazuo Takahashi
摘要简介蛋白质糖基化影响蛋白质的折叠、稳定性、蛋白质相互作用和溶解性等特性。因此，治疗蛋白和可能与疾病发病机制相关的蛋白的糖链部分应该进行深入的分析，包括糖链的异质性和修饰位点。分析方法和仪器的最新进展使高度复杂的糖基化蛋白质的全面表征成为可能。分析糖基化蛋白质时应考虑以下方面：样品制备、层析分离、质谱学(MS)和裂解方法，以及生物信息学，例如用于数据分析的软件解决方案。值得注意的是，分析具有高度唾液酸化的多糖或多个糖基化位点的糖蛋白需要特殊的考虑。在这里，我们讨论了MS的最新方法学进展，提供了异质糖蛋白的详细特征。专家意见由于复杂糖基化蛋白质的特性在分析上仍具有挑战性，这些蛋白质的功能或病理生理学意义尚不完全清楚。为了重复性地生产所需形式的治疗性糖蛋白或充分阐明特定疾病的蛋白质糖基化模式，应该建立一个高度重复性和健壮的分析平台(S)。除了在MS仪器方面的进展外，优化分析和生物信息学方法以及利用糖蛋白/糖肽标准也是可取的。最终，我们设想自动化的高通量MS分析将为临床研究和精确医学提供额外的动力。
ABSTRACT Introduction Protein glycosylation influences characteristics such as folding, stability, protein interactions, and solubility. Therefore, glycan moieties of therapeutic proteins and proteins that are likely associated with disease pathogenesis should be analyzed in-depth, including glycan heterogeneity and modification sites. Recent advances in analytical methods and instrumentation have enabled comprehensive characterization of highly complex glycosylated proteins. Area covered The following aspects should be considered when analyzing glycosylated proteins: sample preparation, chromatographic separation, mass spectrometry (MS) and fragmentation methods, and bioinformatics, such as software solutions for data analyses. Notably, analysis of glycoproteins with heavily sialylated glycans or multiple glycosylation sites requires special considerations. Here, we discuss recent methodological advances in MS that provide detailed characterization of heterogeneous glycoproteins. Expert opinion As characterization of complex glycosylated proteins is still analytically challenging, the function or pathophysiological significance of these proteins is not fully understood. To reproducibly produce desired forms of therapeutic glycoproteins or to fully elucidate disease-specific patterns of protein glycosylation, a highly reproducible and robust analytical platform(s) should be established. In addition to advances in MS instrumentation, optimization of analytical and bioinformatics methods and utilization of glycoprotein/glycopeptide standards is desirable. Ultimately, we envision that an automated high-throughput MS analysis will provide additional power to clinical studies and precision medicine.