Influence of protein (human galectin-3) design on aspects of lectin activity

Abstract

The concept of biomedical significance of the functional pairing between tissue lectins and their glycoconjugate counterreceptors has reached the mainstream of research on the flow of biological information. A major challenge now is to identify the principles of structure–activity relationships that underlie specificity of recognition and the ensuing post-binding processes. Toward this end, we focus on a distinct feature on the side of the lectin, i.e. its architecture to present the carbohydrate recognition domain (CRD). Working with a multifunctional human lectin, i.e. galectin-3, as model, its CRD is used in protein engineering to build variants with different modular assembly. Hereby, it becomes possible to compare activity features of the natural design, i.e. CRD attached to an N-terminal tail, with those of homo- and heterodimers and the tail-free protein. Thermodynamics of binding disaccharides proved full activity of all proteins at very similar affinity. The following glycan array testing revealed maintained preferential contact formation with N-acetyllactosamine oligomers and histo-blood group ABH epitopes irrespective of variant design. The study of carbohydrate-inhibitable binding of the test panel disclosed up to qualitative cell-type-dependent differences in sections of fixed murine epididymis and especially jejunum. By probing topological aspects of binding, the susceptibility to inhibition by a tetravalent glycocluster was markedly different for the wild-type vs the homodimeric variant proteins. The results teach the salient lesson that protein design matters: the type of CRD presentation can have a profound bearing on whether basically suited oligosaccharides, which for example tested positively in an array, will become binding partners in situ. When lectin-glycoconjugate aggregates (lattices) are formed, their structural organization will depend on this parameter. Further testing (ga)lectin variants will thus be instrumental (i) to define the full range of impact of altering protein assembly and (ii) to explain why certain types of design have been favored during the course of evolution, besides opening biomedical perspectives for potential applications of the novel galectin forms.

Glycans in autophagy, endocytosis and lysosomal functions

“…In light of recent findings, other functional oligomeric states of LGALS3 than pentamers, may exist via the dynamic self-association of its intrinsically disordered N-terminal domain [ 156 ]. In a very recent study, Farhadi and colleagues, using engineered LGALS3 constructs (from dimers to hexamers) based on α-helical coiled-coil scaffolds, confirmed that glycan-binding properties, clustering and extracellular signaling activities of LGALS3 depend on architecture and valency, extending previous work with engineered LGALS3 [ 157 – 159 ]. The authors of this study also highlighted that the relatively large integral membrane glycoprotein protein tyrosine phosphatase receptor type C (PTPRC/CD45) (in contrast to smaller glycoproteins such as CD7) regulates membrane glycan clustering and cell death signaling activities of synthetic LGALS3 oligomers [ 157 ].…”

What Cyto- and Histochemistry Can Do to Crack the Sugar Code

“…di- or tetramer) affects staining profiles (and functions). Respective studies have recently been initiated for galectins [ 30 , 60 ].…”

In focus in HCB

“…The pairwise interaction between lectins and their glycoconjugate reaction partners in animal tissues is of importance in many patho-physiological processes (Cummings 2019;Duan and Paulson 2020;Kaltner et al 2019). In their present study, García Caballero et al (2020) aimed to identify principles of structure-activity relationships that underlie specificity of recognition and the ensuing post-binding processes. To this end, they focused on the architecture of the carbohydrate recognition domain (CRD) of the multifunctional human galectin-3 (Gal-3) and engineered variants with different modular assembly to relate protein design to activity.…”