Structural and functional analyses explain Pea KAI2 receptor diversity and reveal stereoselective catalysis during signal perception

Abstract

KAI2 proteins are plant α/β hydrolase receptors which perceive smoke-derived butenolide signals and endogenous, yet unidentified KAI2-ligands (KLs). The number of functional KAI2 receptors varies among species and KAI2 gene duplication and sub-functionalization likely plays an adaptative role by altering specificity towards different KLs. Legumes represent one of the largest families of flowering plants and contain many agronomic crops. Prior to their diversification, KAI2 underwent duplication resulting in KAI2A and KAI2B. Here we demonstrate that Pisum sativum KAI2A and KAI2B are active receptors and enzymes with divergent ligand stereoselectivity. KAI2B has a higher affinity for and hydrolyses a broader range of substrates including strigolactone-like stereoisomers. We determine the crystal structures of PsKAI2B in apo and butenolide-bound states. The biochemical, structural, and mass spectra analyses of KAI2s reveal a transient intermediate on the catalytic serine and a stable adduct on the catalytic histidine, confirming its role as a bona fide enzyme. Our work uncovers the stereoselectivity of ligand perception and catalysis by diverged KAI2 receptors and proposes adaptive sensitivity to KAR/KL and strigolactones by KAI2B.

Nutrient regulation of lipochitooligosaccharide recognition in plants via NSP1 and NSP2

“…Villaécija-Aguilar et al 58 reported that root hair development in d14 is still responsive to 5DS 58 , suggesting a D14 -independent strigolactone receptor. D14L paralogs in pea are able to bind 5DS in vitro and hydrolyse strigolactone-like molecules 59 , raising the possibility that strigolactones or strigolactone-derived molecules 60 may bind and activate D14L.…”

Crystal structure of Arabidopsis DWARF14‐LIKE2 (DLK2) reveals a distinct substrate binding pocket architecture

“…Here, we present the crystal structure of A. thaliana DLK2, which, as a major difference to its paralogs AtD14 and AtKAI2, features a significantly smaller substrate binding pocket. Previous structures of strigolactone receptors or KAI2 clade proteins have shown that not only the substitution of amino acids inside the substrate binding pocket determines its size or ligand specificity (Guercio et al, 2022 ; Toh et al, 2015 ) but that also interactions between residues located in different secondary structure elements can influence the volume and shape of the pocket. This has been observed in a hydrogen bond between helices αD1 and αD3 in hyposensitive to light proteins from Striga hermonthica (ShHTL) (Xu et al, 2018 ) and in a loop region between helices αE and αF in KAI2‐like proteins from Physcomitrium patens (Bürger et al, 2019 ).…”

Strigolactones: diversity, perception, and hydrolysis

“…In general, D14 receptors preferentially perceive SLs where the D ring is in 2 0 R configuration (Fig. 1), whilst KAI2 receptors prefer the 2 0 S configuration (Scaffidi et al 2014;de Saint Germain et al 2016;Carbonnel et al 2020;de Saint Germain et al 2021;Guercio et al 2022). Even within D14 and KAI2 families, these receptors have acquired adaptive sensitivity to different species and/or context-specific SLs, allowing these receptors to represent a wider diversity of ligand specificities.…”