Elucidating the glycan-binding specificity and structure of Cucumis melo agglutinin, a new R-type lectin

Beilstein J Org Chem. 2024 Feb 19:20:306-320. doi: 10.3762/bjoc.20.31. eCollection 2024.


Plant lectins have garnered attention for their roles as laboratory probes and potential therapeutics. Here, we report the discovery and characterization of Cucumis melo agglutinin (CMA1), a new R-type lectin from melon. Our findings reveal CMA1's unique glycan-binding profile, mechanistically explained by its 3D structure, augmenting our understanding of R-type lectins. We expressed CMA1 recombinantly and assessed its binding specificity using multiple glycan arrays, covering 1,046 unique sequences. This resulted in a complex binding profile, strongly preferring C2-substituted, beta-linked galactose (both GalNAc and Fuca1-2Gal), which we contrasted with the established R-type lectin Ricinus communis agglutinin 1 (RCA1). We also report binding of specific glycosaminoglycan subtypes and a general enhancement of binding by sulfation. Further validation using agglutination, thermal shift assays, and surface plasmon resonance confirmed and quantified this binding specificity in solution. Finally, we solved the high-resolution structure of the CMA1 N-terminal domain using X-ray crystallography, supporting our functional findings at the molecular level. Our study provides a comprehensive understanding of CMA1, laying the groundwork for further exploration of its biological and therapeutic potential.

Keywords: R-type; carbohydrate; glycan array; melon; plant lectin.

Grants and funding

This work was funded by a Branco Weiss Fellowship – Society in Science awarded to D.B., by the Knut and Alice Wallenberg Foundation, and the University of Gothenburg, Sweden as well as support from the GLYCONanoPROBES (CA18132) and INNOGLY (CA18103) COST actions awarded to J.L. This work was further supported by the Protein-Glycan Interaction Resource of the CFG and the National Center for Functional Glycomics (NCFG) at Beth Israel Deaconess Medical Center, Harvard Medical School (supporting grant R24 GM137763). The glycan microarray studies were performed in the Carbohydrate Microarray Facility at the ICL Glycosciences Laboratory, which is supported by Wellcome Trust biomedical resource grants (099197/Z/12/Z, 108430/Z/15/Z, and 218304/Z/19/Z) and partially by the March of Dimes Prematurity research centre grant (22-FY18-82). The sequence-defined glycan microarrays contain many saccharides provided by collaborators whom we thank, as well as members of the Glycosciences Laboratory for their contribution in the establishment of the NGL-based microarray system. This work benefited from access to EMBL HTX lab, which has been supported by iNEXT-Discovery, project number 871037, funded by the Horizon 2020 program of the European Commission.