Structural Basis of the Function of Yariv Reagent—An Important Tool to Study Arabinogalactan Proteins

Abstract

Arabinogalactan proteins are very abundant, heavily glycosylated plant cell wall proteins. They are intensively studied because of their crucial role in plant development as well as their function in plant defence. Research of these biomacromolecules is complicated by the lack of tools for their analysis and characterisation due to their extreme heterogeneity. One of the few available tools for detection, isolation, characterisation, and functional studies of arabinogalactan proteins is Yariv reagents. Yariv reagent is a synthetic aromatic glycoconjugate originally prepared as an antigen for immunization. Later, it was found that this compound can precipitate arabinogalactan proteins, namely, their ß-D-(1→3)-galactan structures. Even though this compound has been intensively used for decades, the structural basis of arabinogalactan protein precipitation by Yariv is not known. Multiple biophysical studies have been published, but none of them attempted to elucidate the three-dimensional structure of the Yariv-galactan complex. Here we use a series of molecular dynamics simulations of systems containing one or multiple molecules of ß-D-galactosyl Yariv reagent with or without oligo ß-D-(1→3)-galactan to predict the structure of the complex. According to our model of Yariv-galactan complexes, Yariv reagent forms stacked oligomers stabilized by π-π and CH/π interactions. These oligomers may contain irregularities. Galactan structures crosslink these Yariv oligomers. The results were compared with studies in literature.

The Chirality of Aggregated Yariv Reagents Correlates with Their AGP‐Binding Ability**

“…We speculate that the formation of a helical aggregate is important for binding to an oligogalactan target that itself is postulated to be helical. Support for this hypothesis has been provided by a recent modelling study using small galactan fragments and Yariv reagent oligomers [21] . Shape complementarity between a dye and its molecular target has been observed with DNA‐binding dyes as well as protein‐binding dyes [22] .…”

“…Support for this hypothesis has been provided by a recent modelling study using small galactan fragments and Yariv reagent oligomers. [21] Shape complementarity between a dye and its molecular target has been observed with DNA-binding dyes as well as proteinbinding dyes. [22] Understanding how the widely used Yariv reagents bind to AGP will allow development of better small molecule probes -those that are fluorescent, and bind with high affinity and specificity -to study these important cell wall proteoglycans.…”

O-glycosylation of the extracellular domain of pollen class I formins modulates their plasma membrane mobility

“…The β-Yariv reagent binds selectively to the β-1,3-galactan main chains of AGPs, precipitating them ( Kitazawa et al , 2013 ). β-Yariv is not only used to precipitate AGPs in vitro , but also to perturb AGP function in vivo ( Přerovská et al , 2021 ). Thus, we incubated in vitro- grown pollen tubes with 30 μM β-Yariv and stained the actin cytoskeleton with fluorescent phalloidin.…”

Type II arabinogalactans initiated by hydroxyproline‐O‐galactosyltransferases play important roles in pollen–pistil interactions

“…The Hyp‐ O ‐glycan moieties of AGPs account for more than 90% of their total mass and are suggested to be key players defining their modes of action, based on studies using β‐Yariv reagent (a synthetic phenylglycoside dye that specifically binds to the β‐1,3‐galactan moiety of AGPs) (Kitazawa et al, 2013; Přerovská et al, 2021). Despite the prediction of as many as 85 heterogeneous and complex AGPs in Arabidopsis (Showalter et al, 2010) and extensive studies on several AGPs establishing their crucial roles in reproduction (Coimbra et al, 2007; Pereira, Lopes, & Coimbra, 2016; Pereira, Nobre, et al, 2016), our understanding about the glycan structure–function relationship of AGPs is limited.…”