Publication details
Production of recombinant lectin PluLec from Photorhabdus luminescens for neutron crystallography and its structure-functional characterization.
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| Year of publication | 2023 |
| Type | Appeared in Conference without Proceedings |
| MU Faculty or unit | |
| Citation | |
| Description | Lectins are ubiquitous proteins of non-immune origin that reversibly and specifically interact with carbohydrates. They play a role in various physiological and pathological processes like intercellular communication, adhesion, migration and host-pathogen interactions. Unlike antibodies, they are not products of immune response and lack enzymatic activity. Lectins are widely used for carbohydrate structure characterization, glycoprotein purification, and specific labeling of cell surface structures. Photorhabdus luminescens is a bioluminescent Gram-negative bacterium and an insect pathogen that symbiotically lives in Heterorhabditidae nematodes. PluLec, a putative lectin from Photorhabdus luminescens shares homology with PA-IL lectin, which is D-galactose specific, Ca2+ dependent, cytotoxic lectin from opportunistic pathogen Pseudomonas aeruginosa, involved in facilitating infection in patients with compromised immunity. This research is focused on structural-functional characterization of recombinant protein PluLec using various methods like isothermal titration calorimetry, hemagglutination, glycan array, analytical ultracentrifugation, toxicity tests performed on insect models and protein X-ray and Neutron crystallography. Unraveling lectin’s atomic structure and interactions with carbohydrates is essential for understanding its function and potential applications. Neutron protein crystallography, with its unique ability to visualize hydrogen atom positions, offers a promising avenue for gaining deeper insights into lectin-carbohydrate recognition. However, the successful application of neutron crystallography requires producing sufficient quantities of isotopically labeled recombinant lectin and obtaining high-quality crystals suitable for neutron diffraction. Several considerations, such as selecting an appropriate expression system, optimizing expression and crystallization conditions, need to be taken into account. The study revealed that lectin crystallizes as a homotetramer with four binding sites for D-galactose (one per monomer) and shows specificity towards beta anomers of D-galactose. Preliminary results from toxicity tests conducted on insect models indicate a clear negative effect on insect survival. The obtained results of the structure and function of PluLec may reveal its importance in the pathogenic or symbiotic stage of life. Neutron diffraction experiments can deepen our understanding of lectin functionality, aid in the design of therapeutic strategies targeting lectins, and potentially inspire the development of novel carbohydrate-based therapeutics. |
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