Biochemical and structural studies of target lectin SapL1 from the emerging opportunistic microfungus Scedosporium apiospermum

Abstract

Scedosporium apiospermum is an emerging opportunistic fungal pathogen responsible for life-threatening infections in humans. Host–pathogen interactions often implicate lectins that have become therapeutic targets for the development of carbohydrate mimics for antiadhesive therapy. Here, we present the first report on the identification and characterization of a lectin from S. apiospermum named SapL1. SapL1 was found using bioinformatics as a homolog to the conidial surface lectin FleA from Aspergillus fumigatus known to play a role in the adhesion to host glycoconjugates present in human lung epithelium. In our strategy to obtain recombinant SapL1, we discovered the importance of osmolytes to achieve its expression in soluble form in bacteria. Analysis of glycan arrays indicates specificity for fucosylated oligosaccharides as expected. Submicromolar affinity was measured for fucose using isothermal titration calorimetry. We solved SapL1 crystal structure in complex with α-methyl-L-fucoside and analyzed its structural basis for fucose binding. We finally demonstrated that SapL1 binds to bronchial epithelial cells in a fucose-dependent manner. The information gathered here will contribute to the design and development of glycodrugs targeting SapL1.

Multivalent Fucosides Targeting β-Propeller Lectins from Lung Pathogens with Promising Anti-Adhesive Properties

“…The analysis of the interactions of FleA, SapL1, and BambL with blood group epitopes by glycan array displayed their preference for short, non-branched α-fucosylated oligosaccharides. − The weak affinity for α-methyl fucose, with K d s in the range 1–200 μM, was in some cases ameliorated by its presentation in a disaccharide; especially the α1,3/4 appeared as the preferred fucose linkage and common to all three lectins. For example, the K d of binding for FleA is less than half when passing from the monosaccharide L-Fuc (209 μM) to the disaccharide αFuc(1–4)­GlcNAc (63 μM) in an SPR experiment .…”

“…In our search for novel inhibitors of β-propeller lectins, we employed several biophysical techniques to assess the binding properties of the synthesized compounds. In the expression of FleA and Sapl1, we took advantage of a high-yielding purification method involving a N -terminal tag presenting 6xHis, removed afterward with the Tobacco Etch Virus (TEV) protease to get native protein, as described. , Our attempts to do the same for BambL using the vector pProEX-Htb failed the tag cleavage, presumably due to inaccessibility of the TEV cleavage site. Because of this, the recombinant protein was obtained from the expression of pET25- bambL plasmid, as reported before …”

“…FleA, SapL1, and BambL are β-propeller lectins constituted by antiparallel beta-sheets which organize in six blades around a central funnel-like pore to generate a donut-shaped assembly . Structural characterization by X-ray crystallography showed that SapL1 and FleA share a high similarity in their tertiary and quaternary structures, featuring a homodimeric assembly in which each monomer displays six fucose binding sites at the interface between blades. , BambL, instead, is formed by the trimerization of a shorter peptidic chain . Interestingly, such an arrangement leads to a hexavalent lectin with one binding surface, while the fungal homologues FleA and SapL1 are dodecavalent with two opposite binding faces (Figure ).…”

Structure–function relationship of a novel fucoside-binding fruiting body lectin from Coprinopsis cinerea exhibiting nematotoxic activity

“…Many fucoside-binding lectins from all kingdoms have been characterized. In terms of protein structure, several characterized fucoside-binding lectins from the fungal kingdom adopt a 6-bladed β-propeller fold: Aleuria aurantia agglutinin (AAL) and Aspergillus oryzae lectin (AOL; Wimmerova et al 2003 ; Matsumura et al 2004 ) as well as 2 AOL-homologues that have recently been characterized in the pathogenic molds Aspergillus fumigatus (FleA) and Scedosporium apiospermum (SapL1; Houser et al 2015 ; Martinez-Alarcon et al 2021 ). These lectins are produced mainly by the asexual spores (conidia) of their ascomycete hosts.…”

Novel Treatment Approach for Aspergilloses by Targeting Germination

“…Next to Aspergilli, Scedosporium spp. is the second-most prevalent fungus in the lungs of CF patients [ 240 ] and a homolog of FleA has recently been found and characterized in Scedosporium apiospermum [ 202 ]. Findings of homologs of FleA in other fungi that cause mycosis are promising for the impact potential FleA-targeting antifungals could have on the battle against fungal infections.…”