Structural Basis for Dual Functionality of Isoflavonoid O-Methyltransferases in the Evolution of Plant Defense Responses

Abstract

In leguminous plants such as pea (Pisum sativum), alfalfa (Medicago sativa), barrel medic (Medicago truncatula), and chickpea (Cicer arietinum), 49-O-methylation of isoflavonoid natural products occurs early in the biosynthesis of defense chemicals known as phytoalexins. However, among these four species, only pea catalyzes 3-O-methylation that converts the pterocarpanoid isoflavonoid 6a-hydroxymaackiain to pisatin. In pea, pisatin is important for chemical resistance to the pathogenic fungus Nectria hematococca. While barrel medic does not biosynthesize 6a-hydroxymaackiain, when cell suspension cultures are fed 6a-hydroxymaackiain, they accumulate pisatin. In vitro, hydroxyisoflavanone 49-O-methyltransferase (HI49OMT) from barrel medic exhibits nearly identical steady state kinetic parameters for the 49-O-methylation of the isoflavonoid intermediate 2,7,49-trihydroxyisoflavanone and for the 3-O-methylation of the 6a-hydroxymaackiain isoflavonoid-derived pterocarpanoid intermediate found in pea. Protein x-ray crystal structures of HI49OMT substrate complexes revealed identically bound conformations for the 2S,3R-stereoisomer of 2,7,49-trihydroxyisoflavanone and the 6aR,11aR-stereoisomer of 6a-hydroxymaackiain. These results suggest how similar conformations intrinsic to seemingly distinct chemical substrates allowed leguminous plants to use homologous enzymes for two different biosynthetic reactions. The three-dimensional similarity of natural small molecules represents one explanation for how plants may rapidly recruit enzymes for new biosynthetic reactions in response to changing physiological and ecological pressures.

Structure-Function Analyses of a Caffeic Acid O-Methyltransferase from Perennial Ryegrass Reveal the Molecular Basis for Substrate Preference

“…Lp OMT1 possesses the same tertiary and quaternary structure as several other plant type-1 OMTs that have been characterized crystallographically (Zubieta et al, 2001Liu et al, 2006). Not surprisingly, Lp OMT1 is most structurally similar (a root mean square deviation [rmsd] of 1.51 Å for 332 equivalent residues) to alfalfa COMT , with which it shares 62.7% sequence identity.…”

“…Not surprisingly, Lp OMT1 is most structurally similar (a root mean square deviation [rmsd] of 1.51 Å for 332 equivalent residues) to alfalfa COMT , with which it shares 62.7% sequence identity. Slightly higher rmsd values are observed in comparisons with type-1 OMTs having greater sequence divergence: alfalfa chalcone OMT (Zubieta et al, 2001), 2.19 Å rmsd for 312 equivalent residues, with 40.1% sequence identity; alfalfa isoflavone OMT (Zubieta et al, 2001), 1.91 Å rmsd for 310 equivalent residues, with 28.7% sequence identity; and barrel-medic (Medicago truncatula) isoflavonoid OMT (Liu et al, 2006), 3.15 Å rmsd for 265 equivalent residues, with 19.6% sequence identity.…”

“…For example, in comparison to the structure of (closed-state) isoflavone OMT from alfalfa (Zubieta et al, 2001), the more open conformation of barrel medic isoflavonoid OMT (Liu et al, 2006) was apparent, as evidenced by an outward rotation of the SAM binding domains, a wider active-site cleft, and an inordinately long distance between SAM and the methyl acceptor of the hydroxylated substrate. Similarly, the previously reported structures of ternary complexes of alfalfa COMT …”

Structure and function of enzymes involved in the biosynthesis of phenylpropanoids

“…Chalcone O-methyltransferase (ChOMT) and (iso)flavone O-methyltransferases (IOMTs) use the universal methyl donor S-adenosyl-Lmethionine (SAM) to regiospecifically methylate hydroxyl moieties. Consequently, both enzymes belong to a family of methyltransferases that possess a conserved SAM binding domain α/β Rossman fold consistent with their nucleotide binding activity [43,77]. ChOMT and IOMT (Fig.…”

Negative Regulation of Anthocyanin Biosynthesis in Arabidopsis by a miR156-Targeted SPL Transcription Factor

“…These diphenylchroman compounds play vital roles in growth and development by providing plants with red, blue, and purple pigments that protect against UV radiation, attract pollinators and other beneficial organisms, or mediate plant-microbe interactions (Buer et al, 2010). In addition, many simple flavonoid compounds have antioxidant properties and can therefore potentially be used as dietary nutraceutics for human health (Winkel-Shirley, 2001;Liu et al, 2006). Flavonoid biosynthesis leads to a variety of structurally distinct subclasses, including the flavonols, flavones, isoflavones, anthocyanins, and proanthocyanidins.…”