Wang Wei, Kałuża Anna, Nouta Jan, Nicolardi Simone, Ferens-Sieczkowska Mirosława, Wuhrer Manfred, Lageveen-Kammeijer Guinevere S M, de Haan Noortje
Leiden University Medical Center, Center for Proteomics and Metabolomics, 2300, RC, Leiden, the Netherlands.
Wrocław Medical University, Department of Chemistry and Immunochemistry, Curie-Skłodowska Str. 50, 50-369, Wrocław, Poland.
Talanta. 2021 Jan 15;222:121495. doi: 10.1016/j.talanta.2020.121495. Epub 2020 Aug 13.
An altered total seminal plasma glycosylation has been associated with male infertility, and the highly abundant seminal plasma glycoprotein prostate-specific antigen (PSA) plays an important role in fertilization. However, the exact role of PSA glycosylation in male fertility is not clear. To understand the involvement of PSA glycosylation in the fertilization process, analytical methods are required to study the glycosylation of PSA from seminal plasma with a high glycoform resolution and in a protein-specific manner. In this study, we developed a novel, high-throughput PSA glycopeptide workflow, based on matrix-assisted laser desorption/ionization-mass spectrometry, allowing the discrimination of sialic acid linkage isomers via the derivatization of glycopeptides. The method was successfully applied on a cohort consisting of seminal plasma from infertile and fertile men (N = 102). Forty-four glycopeptides were quantified in all samples, showing mainly complex-type glycans with high levels of fucosylation and sialylation. In addition, N,N-diacetyllactosamine (LacdiNAc) motives were found as well as hybrid-type and high mannose-type structures. Our method showed a high intra- and interday repeatability and revealed no difference in PSA glycosylation between fertile and infertile men. Next to seminal plasma, the method is also expected to be of use for studying PSA glycopeptides derived from other biofluids and/or in other disease contexts.
精液总糖基化改变与男性不育有关，而精液中含量丰富的糖蛋白前列腺特异性抗原（PSA）在受精过程中起重要作用。然而，PSA糖基化在男性生育中的具体作用尚不清楚。为了了解PSA糖基化在受精过程中的作用，需要采用分析方法以高糖型分辨率和蛋白质特异性方式研究精液中PSA的糖基化。在本研究中，我们基于基质辅助激光解吸/电离质谱开发了一种新颖的高通量PSA糖肽工作流程，通过糖肽衍生化实现唾液酸连接异构体的区分。该方法成功应用于一个由不育和生育男性的精液组成的队列（N = 102）。在所有样本中对44种糖肽进行了定量，显示主要为具有高水平岩藻糖基化和唾液酸化的复合型聚糖。此外，还发现了N，N-二乙酰乳糖胺（LacdiNAc）基序以及杂合型和高甘露糖型结构。我们的方法显示出高日内和日间重复性，并且未发现生育和不育男性之间PSA糖基化存在差异。除了精液外，该方法还有望用于研究源自其他生物流体和/或其他疾病背景的PSA糖肽。
