Mucin 1 (MUC1) is a transmembrane mucin expressed at the apical surface of epithelial cells at mucosal surfaces. MUC1 has a barrier function against bacterial invasion and is well known for its aberrant expression and glycosylation in adenocarcinomas. The MUC1 extracellular domain contains a variable number of tandem repeats (VNTR) of 20 amino acids, which are heavily O-linked glycosylated. Monoclonal antibodies against the MUC1 VNTR are powerful research tools with applications in the diagnosis and treatment of MUC1-expressing cancers. Here, we report direct mass spectrometry-based sequencing of anti-MUC1 hybridoma-derived 139H2 IgG, enabling reverse-engineering of the functional recombinant monoclonal antibody. The crystal structure of the 139H2 Fab fragment in complex with the MUC1 epitope was solved, revealing the molecular basis of 139H2 binding specificity to MUC1 and its tolerance to O-glycosylation of the VNTR. The available sequence of 139H2 will allow further development of MUC1-related diagnostic, targeting, and treatment strategies.

中文翻译：

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通过基于质谱的从头测序对抗 MUC1 抗体 139H2 进行逆向工程。

粘蛋白1（MUC1）是一种跨膜粘蛋白，表达于粘膜表面上皮细胞的顶端表面。 MUC1 具有抵抗细菌侵袭的屏障功能，并以其在腺癌中的异常表达和糖基化而闻名。 MUC1 胞外结构域包含可变数量的 20 个氨基酸的串联重复序列 (VNTR)，这些氨基酸均被严重O连接糖基化。针对 MUC1 VNTR 的单克隆抗体是强大的研究工具，可应用于表达 MUC1 的癌症的诊断和治疗。在这里，我们报告了对抗 MUC1 杂交瘤衍生的 139H2 IgG 进行基于质谱的直接测序，从而能够对功能性重组单克隆抗体进行逆向工程。解析了与MUC1表位复合的139H2 Fab片段的晶体结构，揭示了139H2对MUC1的结合特异性及其对VNTR的O-糖基化的耐受性的分子基础。 139H2 的可用序列将允许进一​​步开发 MUC1 相关的诊断、靶向和治疗策略。