Metabolon formation and metabolic channeling in the biosynthesis of plant natural products

Jørgensen, Kirsten; Rasmussen, Anne Vinther; Morant, Marc; Nielsen, Allan Holm; Bjarnholt, Nanna; Zagrobelny, Mika; Bak, Søren; Møller, Birger Lindberg

doi:10.1016/j.pbi.2005.03.014

Cited by 475 publications

(349 citation statements)

References 71 publications

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“…The complex is held together through noncovalent interactions and enables the efficient channeling of a metabolite between two or more active sites, so-called "metabolic channeling" in which processing of metabolic intermediates occurs more quickly than would be expected in a homogeneous aqueous environment. The enzymes of the Calvin and tricarboxylic acid cycles are among the best characterized metabolons (Winkel, 2004;Jørgensen et al, 2005). Metabolic channeling has been demonstrated to occur in the plant phenylpropanoid pathway, and the enzymes of the phenylpropanoid pathway are thought to form such a metabolon, held in place by close associations with membrane-integral cytochrome P450 enzymes (Winkel, 2004;Jørgensen et al, 2005;Lee et al, 2012).…”

Section: Discussionmentioning

confidence: 99%

“…The enzymes of the Calvin and tricarboxylic acid cycles are among the best characterized metabolons (Winkel, 2004;Jørgensen et al, 2005). Metabolic channeling has been demonstrated to occur in the plant phenylpropanoid pathway, and the enzymes of the phenylpropanoid pathway are thought to form such a metabolon, held in place by close associations with membrane-integral cytochrome P450 enzymes (Winkel, 2004;Jørgensen et al, 2005;Lee et al, 2012). Of particular interest here is the suggestion that HCT (also known as CST for hydroxycinnamoylCoA:shikimate hydroxycinnamoyltransferase) and CCoAOMT directly interact in this metabolon, as would be expected of two enzymes catalyzing adjacent steps in the pathway (Winkel, 2004;Jørgensen et al, 2005).…”

Section: Discussionmentioning

confidence: 99%

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Maize Homologs of CCoAOMT and HCT, Two Key Enzymes in Lignin Biosynthesis, Form Complexes with the NLR Rp1 Protein to Modulate the Defense Response

Wang

Balint‐Kurti²

2016

Plant Physiol.

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Disease resistance (R) genes encode nucleotide binding Leu-rich-repeat (NLR) proteins that confer resistance to specific pathogens. Upon pathogen recognition they trigger a defense response that usually includes a so-called hypersensitive response (HR), a rapid localized cell death at the site of pathogen infection. Intragenic recombination between two maize (Zea mays) NLRs, Rp1-D and Rp1-dp2, resulted in the formation of a hybrid NLR, Rp1-D21, which confers an autoactive HR in the absence of pathogen infection. From a previous quantitative trait loci and genome-wide association study, we identified genes encoding two key enzymes in lignin biosynthesis, hydroxycinnamoyltransferase (HCT) and caffeoyl CoA O-methyltransferase (CCoAOMT), adjacent to the nucleotide polymorphisms that were highly associated with variation in the severity of Rp1-D21-induced HR. We have previously shown that the two maize HCT homologs suppress the HR conferred by Rp1-D21 in a heterologous system, very likely through physical interaction. Here, we show, similarly, that CCoAOMT2 suppresses the HR induced by either the full-length or by the N-terminal coiled-coil domain of Rp1-D21 also likely via physical interaction and that the metabolic activity of CCoAOMT2 is unlikely to be necessary for its role in suppressing HR. We also demonstrate that CCoAOMT2, HCTs, and Rp1 proteins can form in the same complexes. A model is derived to explain the roles of CCoAOMT and HCT in Rp1-mediated defense resistance.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Maize Homologs of CCoAOMT and HCT, Two Key Enzymes in Lignin Biosynthesis, Form Complexes with the NLR Rp1 Protein to Modulate the Defense Response

Wang

Balint‐Kurti²

2016

Plant Physiol.

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“…1 and 3). Hydrophobic patches often constitute membrane association or protein interaction sites, and loop B of UGT85B1 may well turn out to serve an important function by mediating attachment of UGT85B1 to CYP79A1 and CYP71E1 to facilitate channeling in the dhurrin pathway by metabolon formation (Jørgensen et al, 2005;Kristensen et al, 2005;Nielsen and Møller, 2005).…”

Section: Discussionmentioning

confidence: 99%

“…In planta, neither p-hydroxymandelonitrile nor its degradation products are detectable, indicating that the glucosylation reaction is highly efficient and channeled (Møller and Conn, 1980). This observation has been substantiated by metabolome and transcriptome analyses of transgenic Arabidopsis (Arabidopsis thaliana) plants expressing the entire dhurrin pathway (Tattersall et al, 2001;Jørgensen et al, 2005;Kristensen et al, 2005), as well as by confocal laser-scanning microscopy studies showing that the three enzymes CYP79A1, CYP71E1, and UGT85B1 catalyzing dhurrin formation are physically associated within a tight metabolon . In contrast, UGT85B1 exhibits a rather broad specificity toward acceptors in vitro, a key determinant for glycosylation being the presence of a sterically unmasked acceptor hydroxyl group (Jones et al, 1999;Hansen et al, 2003).…”

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confidence: 86%

Determination of Catalytic Key Amino Acids and UDP Sugar Donor Specificity of the Cyanohydrin Glycosyltransferase UGT85B1 from Sorghum bicolor. Molecular Modeling Substantiated by Site-Specific Mutagenesis and Biochemical Analyses

Thorsøe¹,

Bak²,

Olsen³

et al. 2005

Plant Physiology

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Plants produce a plethora of structurally diverse natural products. The final step in their biosynthesis is often a glycosylation step catalyzed by a family 1 glycosyltransferase (GT). In biosynthesis of the cyanogenic glucoside dhurrin in Sorghum bicolor, the UDPglucosyltransferase UGT85B1 catalyzes the conversion of p-hydroxymandelonitrile into dhurrin. A structural model of UGT85B1 was built based on hydrophobic cluster analysis and the crystal structures of two bacterial GTs, GtfA and GtfB, which each showed approximately 15% overall amino acid sequence identity to UGT85B1. The model enabled predictions about amino acid residues important for catalysis and sugar donor specificity. p-Hydroxymandelonitrile and UDP-glucose (Glc) were predicted to be positioned within hydrogen-bonding distance to a glutamic acid residue in position 410 facilitating sugar transfer. The acceptor was packed within van der Waals distance to histidine H23. Serine S391 and arginine R201 form hydrogen bonds to the pyrophosphate part of UDP-Glc and hence stabilize binding of the sugar donor. Docking of UDP sugars predicted that UDP-Glc would serve as the sole donor sugar in UGT85B1. This was substantiated by biochemical analyses. The predictive power of the model was validated by site-directed mutagenesis of selected residues and using enzyme assays. The modeling approach has provided a tool to design GTs with new desired substrate specificities for use in biotechnological applications. The modeling identified a hypervariable loop (amino acid residues 156-188) that contained a hydrophobic patch. The involvement of this loop in mediating binding of UGT85B1 to cytochromes P450, CYP79A1, and CYP71E1 within a dhurrin metabolon is discussed.A glycosyltransferase (GT) is an enzyme that attaches a sugar molecule to a specific acceptor and thereby creates a glycosidic bond. GTs are found in all phylae. The nature of the acceptor molecules used is highly diverse, whereas the donor molecules typically are restricted to monosaccharides with either D-or L-configuration linked to a nucleotide (UDP/GDP/ CMP/(d)TDP; Leloir, 1964). Glycosylation reactions constitute an integral part of both primary and secondary metabolism. The final step in plant natural product synthesis is often a glycosylation reaction that serves to stabilize, solubilize, and provide a safe storage form of potentially toxic aglycones. In Sorghum bicolor, the GT UGT85B1 catalyzes glucosylation of Tyr-derived p-hydroxymandelonitrile to yield the cyanogenic glucoside dhurrin (Jones et al., 1999). In planta, neither p-hydroxymandelonitrile nor its degradation products are detectable, indicating that the glucosylation reaction is highly efficient and channeled (Møller and Conn, 1980). This observation has been substantiated by metabolome and transcriptome analyses of transgenic Arabidopsis (Arabidopsis thaliana) plants expressing the entire dhurrin pathway (Tattersall et al., 2001;Jørgensen et al., 2005;Kristensen et al., 2005), as well as by confocal laser-scanning microscopy studies ...

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“…Recently, MdBIS3 was immunochemically localized to the junctions between neighboring cortical parenchyma cells in the transition zone of fire blight-infected apple stems, suggesting an association of the enzyme with plasmodesmata in primary pit fields (Chizzali et al, 2012a. Thus, it is tempting to speculate that the aucuparin biosynthetic enzymes may be assembled into a metabolon, which allows for an efficient transfer of biosynthetic intermediates between the active sites in close proximity (Winkel, 2004;Jørgensen et al, 2005). Protein complexes are commonly associated with structural elements within the cell, such as the ER and microtubules (Zajchowski and Robbins, 2002;Chuong et al, 2004).…”

Section: Resultsmentioning

confidence: 99%

Biphenyl 4-Hydroxylases Involved in Aucuparin Biosynthesis in Rowan and Apple Are Cytochrome P450 736A Proteins

Sircar¹,

Gaid

Chizzali

et al. 2015

Plant Physiol.

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Upon pathogen attack, fruit trees such as apple (Malus spp.) and pear (Pyrus spp.) accumulate biphenyl and dibenzofuran phytoalexins, with aucuparin as a major biphenyl compound. 4-Hydroxylation of the biphenyl scaffold, formed by biphenyl synthase (BIS), is catalyzed by a cytochrome P450 (CYP). The biphenyl 4-hydroxylase (B4H) coding sequence of rowan (Sorbus aucuparia) was isolated and functionally expressed in yeast (Saccharomyces cerevisiae). SaB4H was named CYP736A107. No catalytic function of CYP736 was known previously. SaB4H exhibited absolute specificity for 3-hydroxy-5-methoxybiphenyl. In rowan cell cultures treated with elicitor from the scab fungus, transient increases in the SaB4H, SaBIS, and phenylalanine ammonia lyase transcript levels preceded phytoalexin accumulation. Transient expression of a carboxyl-terminal reporter gene construct directed SaB4H to the endoplasmic reticulum. A construct lacking the amino-terminal leader and transmembrane domain caused cytoplasmic localization. Functional B4H coding sequences were also isolated from two apple (Malus 3 domestica) cultivars. The MdB4Hs were named CYP736A163. When stems of cv Golden Delicious were infected with the fire blight bacterium, highest MdB4H transcript levels were observed in the transition zone. In a phylogenetic tree, the three B4Hs were closest to coniferaldehyde 5-hydroxylases involved in lignin biosynthesis, suggesting a common ancestor. Coniferaldehyde and related compounds were not converted by SaB4H.

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Metabolon formation and metabolic channeling in the biosynthesis of plant natural products

Cited by 475 publications

References 71 publications

Maize Homologs of CCoAOMT and HCT, Two Key Enzymes in Lignin Biosynthesis, Form Complexes with the NLR Rp1 Protein to Modulate the Defense Response

Maize Homologs of CCoAOMT and HCT, Two Key Enzymes in Lignin Biosynthesis, Form Complexes with the NLR Rp1 Protein to Modulate the Defense Response

Determination of Catalytic Key Amino Acids and UDP Sugar Donor Specificity of the Cyanohydrin Glycosyltransferase UGT85B1 from Sorghum bicolor. Molecular Modeling Substantiated by Site-Specific Mutagenesis and Biochemical Analyses

Biphenyl 4-Hydroxylases Involved in Aucuparin Biosynthesis in Rowan and Apple Are Cytochrome P450 736A Proteins

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