Podoplanin (PDPN) is a highly O-glycosylated glycoprotein that is utilized as a specific lymphatic endothelial marker under pathophysiological conditions. We previously developed an anti-human PDPN (hPDPN) monoclonal antibody (mAb), clone LpMab-3, which recognizes the epitope, including both the peptides and the attached disialy-core-l (NeuAcα2-3Galβl-3 [NeuAcα2-6]GalNAcαl-O-Thr) structure at the Thr76 residue in hPDPN. However, it is unclear if the mAb binds directly to both the peptides and glycans. In this study, we synthesized the binding epitope region of LpMab-3 that includes the peptide (-₆₇LVATSVNSV-T-GIRIEDLP₈₄-) possessing a disialyl-core-1 O-glycan at Thr76, and we determined the crystal structure of the LpMab-3 Fab fragment that was bound to the synthesized glycopeptide at a 2.8 Å resolution. The six amino acid residues and two sialic acid residues are directly associated with four complementarity-determining regions (CDRs; H1, H2, H3, and L3) and four CDRs (H2, H3, L1, and L3), respectively. These results suggest that IgG is advantageous for generating binders against spacious epitopes such as glycoconjugates.

Podoplanin（PDPN）是一种高度O-糖基化的糖蛋白，在病理生理条件下可用作特定的淋巴管内皮标记。我们之前开发了抗人PDPN（hPDPN）单克隆抗体（mAb），克隆LpMab-3，可识别抗原决定簇，包括肽和附着的二核核心-1（NeuAcα2-3Galβ1-3[NeuAcα2-6]在hPDPN中的Thr76残基处的GalNAcα1 - O- Thr）结构。然而，尚不清楚mAb是否直接结合到肽和聚糖上。在这项研究中，我们合成LpMab-3的结合表位区，其包括所述肽（ - ₆₇ LVATSVNSV-T-GIRIEDLP ₈₄具有一个disialyl-核心-1 - ）Ö-Thr76处的β-聚糖，我们确定了LpMab-3 Fab片段的晶体结构，该片段以2.8Å的分辨率与合成的糖肽结合。六个氨基酸残基和两个唾液酸残基分别直接与四个互补决定区（CDR； H1，H2，H3和L3）和四个CDR（H2，H3，L1和L3）关联。这些结果表明，IgG对于产生针对宽表位例如糖缀合物的结合剂是有利的。