Functional analysis of members of the isoflavone and isoflavanone O-methyltransferase enzyme families from the model legume Medicago truncatula

Abstract

Previous studies have identified two distinct O-methyltransferases (OMTs) implicated in isoflavonoid biosynthesis in Medicago species, a 7-OMT methylating the A-ring 7-hydroxyl of the isoflavone daidzein and a 4'-OMT methylating the B-ring 4'-hydroxyl of 2,7,4'-trihydroxyisoflavanone. Genes related to these OMTs from the model legume Medicago truncatula cluster as separate branches of the type I plant small molecule OMT family. To better understand the possible functions of these related OMTs in secondary metabolism in M. truncatula, seven of the OMTs were expressed in E. coli, purified, and their in vitro substrate preferences determined. Many of the enzymes display promiscuous activities, and some exhibit dual regio-specificity for the 4' and 7-hydroxyl moieties of the isoflavonoid nucleus. Protein structure homology modeling was used to help rationalize these catalytic activities. Transcripts encoding the different OMT genes exhibited differential tissue-specific and infection- or elicitor-induced expression, but not always in parallel with changes in expression of confirmed genes of the isoflavonoid pathway. The results are discussed in relation to the potential in vivo functions of these OMTs based on our current understanding of the phytochemistry of M. truncatula, and the difficulties associated with gene annotation in plant secondary metabolism.

Two O-methyltransferases involved in the biosynthesis of methoxypyrazines: grape-derived aroma compounds important to wine flavour

“…2b) suggesting that a possible duplication event followed by mutations may have led to the divergent evolution of the two genes. A similar occurrence has been reported in the legume Medicago truncatula in which three OMTs with different substrate specificities are located within a 20 kb region of the genome (Deavours et al 2006). Truncated gene fragments with homology to methyltransferases were located between VvOMT1 and VvOMT2, although no ESTs matching these sequences were found in the public databases.…”

Metabolomics Reveals Novel Pathways and Differential Mechanistic and Elicitor-Specific Responses in Phenylpropanoid and Isoflavonoid Biosynthesis in Medicago truncatula Cell Cultures

“…The biogenetic origins of many of these compounds have been determined from previous studies with chickpea (Cicer arietinum) and alfalfa (Edwards and Kessman, 1992;Dixon, 1999). However, three methylated isoflavones, afrormosin, alfalone, and irisolidone, were identified here for the first time in M. truncatula, and the biosynthetic origins of these and other methylated isoflavonoids is still a matter of some uncertainty (Edwards and Dixon, 1991;Deavours et al, 2006).…”

Functional Analysis of a Predicted Flavonol Synthase Gene Family in Arabidopsis

“…Therefore, it appears that the gene family model described for FLS, with one catalytically active member and several pseudogenes, may also apply to other flavonoid genes, particularly in the case of CHI. There is also a growing awareness that the ''promiscuity'' of metabolic enzymes such as ANS, as well as flavonoid glycosyltransferases (Lim et al, 2004) and O-methyltransferases (Deavours et al, 2006), is more the rule than the exception (Taglieber et al, 2007). These ''alternative'' functions of otherwise well-characterized proteins may represent new paradigms that must be taken into account in efforts to develop framework models of cellular metabolism.…”