Lectin-Binding Specificity of the Fertilization-Relevant Protein PDC-109 by Means of Surface Plasmon Resonance and Carbohydrate REcognition Domain EXcision-Mass Spectrometry

Sira Defaus,Ricardo Gutiérrez-Gallego,David Andreu,Manuel Avilés

doi:10.3390/ijms19041076

Abstract

Seminal plasma proteins are relevant for sperm functionality and some appear responsible for establishing sperm interactions with the various environments along the female genital tract towards the oocyte. In recent years, research has focused on characterizing the role of these proteins in the context of reproductive biology, fertility diagnostics and treatment of related problems. Herein, we focus on the main protein of bovine seminal plasma, PDC-109 (BSP-A1/-A2), which by virtue of its lectin properties is involved in fertilization. By means of surface plasmon resonance, the interaction of PDC-109 with a panel of the most relevant glycosidic epitopes of mammals has been qualitatively and quantitatively characterized, and a higher affinity for carbohydrates containing fucose has been observed, in line with previous studies. Additionally, using the orthogonal technique of Carbohydrate REcognition Domain EXcision-Mass Spectrometry (CREDEX-MS), the recognition domain of the interaction complexes between PDC-109 and all fucosylated disaccharides [(Fuc-α1,(3,4,6)-GlcNAc)] has been defined, revealing the specific glycotope and the peptide domain likely to act as the PDC-109 carbohydrate binding site.

Highlights

In the male genital tract, secretions from the testes, epididymis, seminal vesicles, and other accessory glands contribute to the fluid portion of semen, in which sperm cells are suspended
We aimed to study the lectin-binding profile of PDC-109 using purified protein material from biological origin, initial work was focused on the synthesis, purification and validation of new glycoprobes displaying some of the most relevant glycotopes in the mammalian system (Table 1)
Ligation reactions were performed at pH 3.5 (NAc-hexoses) or pH 4.6 and 37 ◦C for 72 h, the conjugates purified by high performance liquid chromatography (HPLC) and characterized by mass spectrometry (MS) as detailed in Materials and Methods

Summary

Introduction

In the male genital tract, secretions from the testes, epididymis, seminal vesicles, and other accessory glands contribute to the fluid portion of semen (i.e., seminal plasma), in which sperm cells (i.e., spermatozoa) are suspended. Since sperm cells have a limited biosynthetic ability [1], the interaction of different seminal plasma proteins with spermatozoa, as well as with proteins from the female genital tract encountered during the journey to the site of fertilization, will have a profound impact on sperm functionality. Studies on the nature, structure, and functional properties of seminal plasma protein constituents, and on their interactions with molecular ligands and spermatozoa, might provide clues on how seminal plasma modulates the functional ability of spermatozoa. These insights are likely to help advance aspects of such as diagnostics and treatment of reproductive dysfunction, as semen reflects the status of the male reproductive organs

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Conclusion