Characterization ofS-Adenosyl-l-Methionine:(Iso)eugenolO-Methyltransferase Involved in Floral Scent Production inClarkia breweri

Wang, Jihong; Pichersky, Eran

doi:10.1006/abbi.1997.0452

Cited by 76 publications

(60 citation statements)

References 14 publications

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“…Site-saturation Mutagenesis-The cDNA encoding C. breweri IEMT was PCR amplified from the pET11-IEMT plasmid (24) and subcloned into a modified pET28a(ϩ) vector compatible with Gateway cloning to fuse the IEMT with a His tag to facilitate Ni ϩ -mediated affinity protein purification (28). Saturation mutagenesis was performed at sites 130, 131, 133, 134,139,164,165, 175,186, 319, 326, and 327 of IEMT following the QuikChange site-directed mutagenesis strategy (Stratagene) using NNK degenerate primers (N represents a mixture of A, T, G, C, and K for G/T) (supplemental Table S1).…”

Section: Chemicals-[methyl-mentioning

confidence: 99%

“…In Clarkia breweri (fairy fans) and sweet basil (Ocimum basilicum), the identified O-methyltransferases catalyze the 4-O-methylation of a group of volatile compounds, viz. the phenylpropenes, isoeugenol, eugenol, and/or chavicol (23,24). These allyl/propenylphenols are structural analogs of monolignols, differing only in their propanoid tail.…”

mentioning

confidence: 99%

“…1A). Early pioneering studies by Wang and Pichersky (19,24) revealed that (iso)eugenol 4-O-methyltransferase (IEMT) (EC. 2.1.1.146) evolved from COMT by gene duplication and subsequent mutations.…”

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confidence: 99%

“…1A). Swapping sequence fragments of IEMT and COMT, or reversely converting the residues around their active sites intertransformed their activities, although it greatly compromised the catalytic efficiency of the resulting enzyme variants (19,24). Interestingly, these designed site-directed mutations invariably and simultaneously alter substrate preference and regioselective methylation, i.e.…”

mentioning

confidence: 99%

“…Interestingly, these designed site-directed mutations invariably and simultaneously alter substrate preference and regioselective methylation, i.e. the mutations change the activity of IEMT from the 4-O-methylation of allyl/propenylphenols to the 3/5-O-methylation of caffeic acid, and vice versa (19,24).…”

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confidence: 99%

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Engineering Monolignol 4-O-Methyltransferases to Modulate Lignin Biosynthesis

Bhuiya

Liu

2010

Journal of Biological Chemistry

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Lignin is a complex polymer derived from the oxidative coupling of three classical monolignols. Lignin precursors are methylated exclusively at the meta-positions (i.e. 3/5-OH) of their phenyl rings by native O-methyltransferases, and are precluded from substitution of the para-hydroxyl (4-OH) position. Ostensibly, the para-hydroxyls of phenolics are critically important for oxidative coupling of phenoxy radicals to form polymers. Therefore, creating a 4-O-methyltransferase to substitute the para-hydroxyl of monolignols might well interfere with the synthesis of lignin. The phylogeny of plant phenolic O-methyltransferases points to the existence of a batch of evolutionarily "plastic" amino acid residues. Following one amino acid at a time path of directed evolution, and using the strategy of structure-based iterative site-saturation mutagenesis, we created a novel monolignol 4-O-methyltransferase from the enzyme responsible for methylating phenylpropenes. We show that two plastic residues in the active site of the parental enzyme are vital in dominating substrate discrimination. Mutations at either one of these separate the evolutionarily tightly linked properties of substrate specificity and regioselective methylation of native O-methyltransferase, thereby conferring the ability for para-methylation of the lignin monomeric precursors, primarily monolignols. Beneficial mutations at both sites have an additive effect. By further optimizing enzyme activity, we generated a triple mutant variant that may structurally constitute a novel phenolic substrate binding pocket, leading to its high binding affinity and catalytic efficiency on monolignols. The 4-O-methoxylation of monolignol efficiently impairs oxidative radical coupling in vitro, highlighting the potential for applying this novel enzyme in managing lignin polymerization in planta.

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Section: Chemicals-[methyl-mentioning

confidence: 99%

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confidence: 99%

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confidence: 99%

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confidence: 99%

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Engineering Monolignol 4-O-Methyltransferases to Modulate Lignin Biosynthesis

Bhuiya

Liu

2010

Journal of Biological Chemistry

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Biogenesis of Floral Scents

Dudareva

Piechulla

Pichersky

1999

Horticultural Reviews

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Biosynthesis of Phenylpropanoids and Related Compounds (FromAPRVolume 40)

Petersen

Hans

Matern

2018

Annual Plant Reviews Online

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Phenolic compounds are ubiquitous in the plant kingdom. A main pathway for the formation of these compounds starts with the aromatic amino acidsl‐phenylalanine and – to a lesser extent –l‐tyrosine. In the general phenylpropanoid pathway, these are transformed to coenzyme A‐activated 4‐coumaric acid by phenylalanine ammonia‐lyase, cinnamic acid 4‐hydroxylase and 4‐coumarate CoA‐ligase. 4‐Coumaroyl‐CoA gives rise to a large number of different natural products, e.g. flavonoids, lignans, coumarins, tannins, hydroxycinnamic acid esters and amides as well as lignin monomers. This review gives an insight into recent findings concerning the general phenylpropanoid pathway as well as the biosyntheses of hydroxycinnamoyl conjugates, phenolic aroma and fragrance compounds, lignans, coumarins and gallo‐/ellagitannins.

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Characterization ofS-Adenosyl-l-Methionine:(Iso)eugenolO-Methyltransferase Involved in Floral Scent Production inClarkia breweri

Cited by 76 publications

References 14 publications

Engineering Monolignol 4-O-Methyltransferases to Modulate Lignin Biosynthesis

Engineering Monolignol 4-O-Methyltransferases to Modulate Lignin Biosynthesis

Biogenesis of Floral Scents

Biosynthesis of Phenylpropanoids and Related Compounds (FromAPRVolume 40)

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