Metabolite Channeling and Multi-enzyme Complexes



The assembly of cooperating enzymes into multicatalytic complexes, also known as “metabolons,” has become a well-accepted concept in cellular metabolism, at least in principle. There are still relatively few examples where the existence of such systems is supported by solid experimental evidence and even fewer where there is evidence for “channeling” of metabolites through the complex. However, proteomic approaches are providing new evidence for the pervasiveness of this type of organization, while structural biology is offering insights into how these systems are constructed and regulated. New and improved technologies for analyzing protein interactions and assemblies, both in vitro and in intact cells, are opening the doors to explo-ring the intracellular organization of a growing number of metabolic complexes in plants and other organisms. There is also an increasing appre-ciation of the surprising scale of many protein interaction networks, the multiple functions of individual proteins, and the importance (and challenges) of compartmentalization. As a result, the concept of enzyme complexes is gaining wider acceptance and becoming an increasingly important consideration in efforts to engineer metabolism.


Protein Interaction Network Multienzyme Complex Cysteine Biosynthesis Multienzyme System Metabolic Channeling 



The author acknowledges the insights of Joe Chappel (on the erg28p system), Danny Kohl (on the implications of recent metabolic profiling experiments in E. coli), and Joe Noel (on the issue of enzyme promiscuity supporting the existence of enzyme complexes), as well as the very helpful comments of three anonymous reviewers. She is grateful to the National Science Foundation for supporting the work in her laboratory on the flavonoid enzyme complex (currently grant number MCB 0445878). This article is dedicated to the memory of H. Olin Spivey, Professor Emeritus of Biochemistry and Molecular Biology at Oklahoma State University, who died on December 5, 2007. A pioneer in the field of metabolic organization, he will be remembered by many as a valued colleague who encouraged newcomers to join the network.


  1. 1.
    Jorgensen, al (2005) Metabolon formation and metabolic channeling in the biosynthesis of plant natural products. Curr. Opin. Plant Biol.8, 280–291PubMedCrossRefGoogle Scholar
  2. 2.
    Marsh, al (2001) Organellar relationships in the Golgi region of the pancreatic beta cell line, HIT-T15, visualized by high resolution electron tomography. Proc. Natl. Acad. Sci. USA.98, 2399–2406PubMedCrossRefGoogle Scholar
  3. 3.
    Uetz, al (2000) A comprehensive analysis of protein-protein interactions in Saccharomyces cerevisiae<. Nature403, 623–627PubMedCrossRefGoogle Scholar
  4. 4.
    Giot, al (2003) A protein interaction map of Drosophila melanogaster<. Science302, 1727–1736PubMedCrossRefGoogle Scholar
  5. 5.
    Rual, al (2005) Towards a proteome-scale map of the human protein-protein interaction network. Nature437, 1173–1178PubMedCrossRefGoogle Scholar
  6. 6.
    Li, al (2004) A map of the interactome network of the metazoan C. elegans. Science303, 540–543PubMedCrossRefGoogle Scholar
  7. 7.
    Han, al (2004) Evidence for dynamically organized modularity in the yeast protein-protein interaction network. Nature430, 88–93PubMedCrossRefGoogle Scholar
  8. 8.
    Tarassov, al (2008) An in vivo map of the yeast protein interactome. Science320, 1465–1470PubMedCrossRefGoogle Scholar
  9. 9.
    Robinson, al (2007) The molecular sociology of the cell. Nature450, 973–982PubMedCrossRefGoogle Scholar
  10. 10.
    Srere, P.A. (2000) Macromolecular interactions: tracing the roots. Trends Biochem. Sci.25, 150–153PubMedCrossRefGoogle Scholar
  11. 11.
    Islam, al (2007) A novel branched-chain amino acid metabolon – protein-protein interactions in a supramolecular complex. J. Biol. Chem.282, 11893–11903PubMedCrossRefGoogle Scholar
  12. 12.
    Ishikawa, al (2004) Structural basis for channelling mechanism of a fatty acid beta-oxidation multienzyme complex. EMBO J.23, 2745–2754PubMedCrossRefGoogle Scholar
  13. 13.
    Jenni, al (2006) Architecture of a fungal fatty acid synthase at 5 angstrom resolution. Science311, 1263–1267PubMedCrossRefGoogle Scholar
  14. 14.
    Maier, al (2006) Architecture of mammalian fatty acid synthase at 4.5 angstrom resolution. Science311, 1258–1262PubMedCrossRefGoogle Scholar
  15. 15.
    Fries, al (2007) Distinct modes of recognition of the lipoyl domain as substrate by the E1 and E3 components of the pyruvate dehydrogenase multienzyme complex. J. Mol. Biol.366, 132–139PubMedCrossRefGoogle Scholar
  16. 16.
    Giles, N.H. (1978) The organization, function, and evolution of gene clusters in eucaryotes. Am. Naturalist112, 641–657CrossRefGoogle Scholar
  17. 17.
    Singh, S.A. and Christendat, D. (2007) The DHQ-dehydroshikimate-SDH-shikimate-NADP(H) complex: insights into metabolite transfer in the shikimate pathway. Cryst. Growth Des.7, 2153–2160CrossRefGoogle Scholar
  18. 18.
    Luo, al (2007) Simultaneous determination of multiple intracellular metabolites in glycolysis, pentose phosphate pathway and tricarboxylic acid cycle by liquid chromatography-mass spectro-metry. J. Chromatogr. A1147, 153–164PubMedCrossRefGoogle Scholar
  19. 19.
    Ishii, al (2007) Multiple high-throughput analyses monitor the response of E. coli to perturbations. Science316, 593–597PubMedCrossRefGoogle Scholar
  20. 20.
    Saigne-Soulard, al (2006) C-13 NMR analysis of polyphenol biosynthesis in grape cells: Impact of various inducing factors. Anal. Chim. Acta563, 137–144CrossRefGoogle Scholar
  21. 21.
    Pereira, M.P. and Brown, E.D. (2004) Bifunctional catalysis by CDP-ribitol synthase: Convergent recruitment of reductase and cytidylyltransferase activities in Haemophilus influenzae< and Staphylococcus aureus<. Biochemistry (Mosc).43, 11802–11812CrossRefGoogle Scholar
  22. 22.
    Garcon, al (2006) Crystal structure of the bifunctional dihydroneopterin aldolase/6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase from Streptococcus pneumoniae<. J. Mol. Biol.360, 644–653PubMedCrossRefGoogle Scholar
  23. 23.
    Taglieber, A. et al (2007) Alternate-site enzyme promiscuity. Angew. Chem. Int. Ed.46, 8597–8600CrossRefGoogle Scholar
  24. 24.
    Peisajovich, S.G. and Tawfik, D.S. (2007) Protein engineers turned evolutionists. Nature Methods4, 991–994PubMedCrossRefGoogle Scholar
  25. 25.
    Chiron, al (2000) Molecular cloning and functional expression of a stress-induced multifunctional O-methyltransferase with pinosylvin methyltransferase activity from Scots pine (Pinus sylvestris< L.). Plant Mol. Biol.44, 733–745PubMedCrossRefGoogle Scholar
  26. 26.
    Frick, S. and Kutchan, T.M. (1999) Molecular cloning and functional expression of O<-methyl-transferases common to isoquinoline alkaloid and phenylpropanoid biosynthesis. Plant J.17, 329–339PubMedCrossRefGoogle Scholar
  27. 27.
    Gauthier, al (1998) Characterization of two cDNA clones which encode O<-methyl­transferases for the methylation of both flavonoid and phenylpropanoid compounds.Arch. Biochem. Biophys.351, 243–249PubMedCrossRefGoogle Scholar
  28. 28.
    He, X.-Z. and Dixon, R.A. (2000) Genetic mani-pulation of isoflavone 7-O-methyltransferase enhances biosynthesis of 4’-O-methylated isoflavonoid phytoalexins and disease resistance in alfalfa. Plant Cell12, 1689–1702PubMedCrossRefGoogle Scholar
  29. 29.
    Liu, C.-J. and Dixon, R.A. (2001) Elicitor-induced association of isoflavone O-methyltransferase with endomembranes prevents the formation and 7-O-methylation of daizein during isoflavoniod phytoalexin biosynthesis. Plant Cell13, 2643–2658PubMedCrossRefGoogle Scholar
  30. 30.
    Deavours, B.E. et al (2006) Functional analysis of members of the isoflavone and isoflavanone O-methyltransferase enzyme families from the model legume Medicago truncatula. Plant Mol. Biol.62, 715–733PubMedCrossRefGoogle Scholar
  31. 31.
    Liu, C.J. et al (2006) Structural basis for dual functionality of isoflavonoid O-methyl­transferases in the evolution of plant defense responses. Plant Cell18, 3656–3669PubMedCrossRefGoogle Scholar
  32. 32.
    Zubieta, C. et al (2001) Structures of two natural product methyltransferases reveal the basis for substrate specificity in plant O-methyltransferases. Nat. Struct. Biol.8, 271–279PubMedCrossRefGoogle Scholar
  33. 33.
    Jenrich, al (2007) Evolution of heteromeric nitrilase complexes in Poaceae with new functions in nitrile metabolism. Proc. Natl. Acad. Sci. USA104, 18848–18853PubMedCrossRefGoogle Scholar
  34. 34.
    Kriechbaumer, al (2007) Maize nitrilases have a dual role in auxin homeostasis and β-cyanoalanine hydrolysis. J. Exp. Bot.58, 4225–4233PubMedCrossRefGoogle Scholar
  35. 35.
    Kim, al (2005) Novel type of enzyme multimerization enhances substrate affinity of oat β-glucosidase. J. Struct. Biol.150, 1–10PubMedCrossRefGoogle Scholar
  36. 36.
    Owens, al (2008) Functional analysis of a predicted flavonol synthase gene family in Arabidopsis. Plant Physiol.147, 1046–1061PubMedCrossRefGoogle Scholar
  37. 37.
    Kim, al (2004) Functional reclassification of the putative cinnamyl alcohol dehydrogenase multigene family in Arabidopsis. Proc. Natl. Acad. Sci. USA101, 1455–1460PubMedCrossRefGoogle Scholar
  38. 38.
    Yamagami, T. et al (2003) Biochemical diversity among the 1-amino-cyclopropane-1-carboxylate synthase isozymes encoded by the Arabidopsis gene familyJ. Biol. Chem.278, 49102–49112PubMedCrossRefGoogle Scholar
  39. 39.
    Dhugga, K.S. (2007) Maize biomass yield and composition for biofuels. Crop Sci.47, 2211–2227CrossRefGoogle Scholar
  40. 40.
    Somerville, C. (2006) Cellulose synthesis in higher plants. Annu. Rev. Cell Dev. Biol.22, 53–78PubMedCrossRefGoogle Scholar
  41. 41.
    Saxena, I.M. and Brown, R.M. (2005) Cellulose biosynthesis: current views and evolving concepts. Ann. Bot.96, 9–21PubMedCrossRefGoogle Scholar
  42. 42.
    Mueller, S.C. and Brown, R.M. (1980) Evidence for an intramembrane component associated with a cellulose microfibril-synthesizing complex in higher plants. J. Cell Biol.84, 315–326PubMedCrossRefGoogle Scholar
  43. 43.
    Taylor, al (2003) Interactions among three distinct CesA proteins essential for cellulose synthesis. Proc. Natl. Acad. Sci. USA100, 1450–1455PubMedCrossRefGoogle Scholar
  44. 44.
    Persson, S. et al (2007) Genetic evidence for three unique components in primary cell-wall cellulose synthase complexes in Arabidopsis<. Proc. Natl. Acad. Sci. USA104, 15566–15571PubMedCrossRefGoogle Scholar
  45. 45.
    Desprez, T. et al (2007) Organization of cellulose synthase complexes involved in primary cell wall synthesis in Arabidopsis thaliana<. Proc. Natl. Acad. Sci. USA104, 15572–15577PubMedCrossRefGoogle Scholar
  46. 46.
    Paradez, al (2006) Microtubule cortical array organization and plant cell morphogenesis. Curr. Opin. Plant Biol.9, 571–578PubMedCrossRefGoogle Scholar
  47. 47.
    DeBolt, al (2007) Morlin, an inhibitor of cortical microtubule dynamics and cellulose synthase movement. Proc. Natl. Acad. Sci. USA104, 5854–5859PubMedCrossRefGoogle Scholar
  48. 48.
    Chuong, al (2004) Large-scale identification of tubulin-binding proteins provides insight on subcellular trafficking, metabolic channeling, and signaling in plant cells. Mol. Cell. Proteomics3, 970–983PubMedCrossRefGoogle Scholar
  49. 49.
    Hrazdina, G. and Wagner, G.J. (1985) Metabolic pathways as enzyme complexes: evidence for the synthesis of phenylpropanoids and flavonoids on membrane associated enzyme complexes. Arch. Biochem. Biophys.237, 88–100PubMedCrossRefGoogle Scholar
  50. 50.
    Hughes, al (2000) Functional discovery via a compendium of expression profiles. Cell102, 109–126PubMedCrossRefGoogle Scholar
  51. 51.
    Mo, C. and Bard, M. (2005) Erg28p is a key protein in the yeast sterol biosynthetic enzyme complex. J. Lipid Res.46, 1991–1998PubMedCrossRefGoogle Scholar
  52. 52.
    Mo, C. and Bard, M. (2005) A systematic study of yeast sterol biosynthetic protein-protein interactions using the split-ubiquitin system. Biochim. Biophys. Acta Mol. Cell. Biol. Lipids1737, 152–160Google Scholar
  53. 53.
    Ottolenghi, al (2000) The genomic structure of C140rf1 is conserved across eukarya. Mamm. Genome11, 786–788PubMedCrossRefGoogle Scholar
  54. 54.
    Burbulis, I.E. and Winkel-Shirley, B. (1999) Interactions among enzymes of the Arabidopsis< flavonoid biosynthetic pathway. Proc. Natl. Acad. Sci. USA96, 12929–12934PubMedCrossRefGoogle Scholar
  55. 55.
    Owens, D.K. et al. (2008) Biochemical and genetic characterization of Arabidopsis flavanone 3β-hydroxylase. Plant Physiol. Biochem. 46, 833–843Google Scholar
  56. 56.
    Kredich, N. et al (1969) Purification and characterization of cysteine synthetase, a bifunctional protein complex, from Salmonella typhimurium<. J. Biol. Chem.244, 2428–2439PubMedGoogle Scholar
  57. 57.
    Wirtz, M. and Hell, R. (2006) Functional analysis of the cysteine synthase protein complex from plants: Structural, biochemical and regulatory properties. J. Plant Physiol.163, 273–286PubMedCrossRefGoogle Scholar
  58. 58.
    Bonner, al (2005) Molecular basis of cysteine biosynthesis in plants – structural and functional analysis of O-acetylserine sulfhydrylase from Arabidopsis thaliana<. J. Biol. Chem.280, 38803–38813PubMedCrossRefGoogle Scholar
  59. 59.
    Wirtz, M. and Hell, R. (2007) Dominant-negative modification reveals the regulatory function of the multimeric cysteine synthase protein complex in transgenic tobacco. Plant Cell19, 625–639PubMedCrossRefGoogle Scholar
  60. 60.
    Francois, J.A. et al (2006) Structural basis for interaction of O-acetylserine sulfhydrylase and serine acetyltransferase in the Arabidopsis< cyst-eine synthase complex. Plant Cell18, 3647–3655PubMedCrossRefGoogle Scholar
  61. 61.
    Kumaran, S. and Jez, J.M. (2007) Ther-modynamics of the interaction between O-acetylserine sulfhydrylase and the C-terminus of serine acetyltransferase. Biochemistry (Mosc).46, 5586–5594CrossRefGoogle Scholar
  62. 62.
    Petoukhov, M.V. and Svergun, D.I. (2007) Analysis of X-ray and neutron scattering from biomacromolecular solutions. Curr. Opin. Struct. Biol.17, 562–571PubMedCrossRefGoogle Scholar
  63. 63.
    Anderson, L.E. and Carol, A.A. (2005) Enzyme co-localization in the pea leaf cytosol: 3-P-glycerate kinase, glyceraldehyde-3-P dehydrogenase, triose-P isomerase and aldolase. Plant Sci.169, 620–628CrossRefGoogle Scholar
  64. 64.
    Graham, al (2007) Glycolytic enzymes associate dynamically with mitochondria in response to respiratory demand and support substrate channeling. Plant Cell19, 3723–3738PubMedCrossRefGoogle Scholar
  65. 65.
    Dudkina, N.V. et al (2006) Respiratory chain supercomplexes in the plant mitochondrial membrane. Trends Plant Sci.11, 232–240PubMedCrossRefGoogle Scholar
  66. 66.
    Winkel, B. (2004) Metabolic channeling in plants. Annu. Rev. Plant Biol.55, 85–107PubMedCrossRefGoogle Scholar
  67. 67.
    Facchini, P.J. and St-Pierre, B. (2005) Synthesis and trafficking of alkaloid biosynthetic enzymes. Curr. Opin. Plant Biol.8, 657–666PubMedCrossRefGoogle Scholar
  68. 68.
    Gómez-Galera, al (2007) The genetic manipulation of medicinal and aromatic plants. Plant Cell Rep.26, 1689–1715PubMedCrossRefGoogle Scholar
  69. 69.
    Morant, A.V. et al (2007) Lessons learned from metabolic engineering of cyanogenic glucosides. Metabolomics3, 383–398CrossRefGoogle Scholar
  70. 70.
    Shimamura, al (2007) 2-hydroxyisoflavanone dehydratase is a critical determinant of isoflavone productivity in hairy root cultures of Lotus japo-nicus. Plant Cell Physiol.48, 1652–1657PubMedCrossRefGoogle Scholar
  71. 71.
    Aharoni, A. et al (2005) Volatile science? Metabolic engineering of terpenoids in plants. Trends Plant Sci.10, 594–602PubMedCrossRefGoogle Scholar
  72. 72.
    Yan, al (2005) Metabolic engineering of anthocyanin biosynthesis in Escherichia coli<. Appl. Environ. Microbiol.71, 3617–3623PubMedCrossRefGoogle Scholar
  73. 73.
    Zalokar, M. (1960) Cytochemistry of centrifuged hyphae of Neurospora<. Exp. Cell Res.19, 114–132PubMedCrossRefGoogle Scholar
  74. 74.
    Zalokar, M. (1969) Intracellular centrifugal separation of organelles in Phycomyces<. J. Cell Biol.41, 494–509PubMedCrossRefGoogle Scholar
  75. 75.
    Kempner, E.S. and Miller, J.H. (1968) The molecular biology of Euglena gracilis. Exp. Cell Res.51, 141–149PubMedCrossRefGoogle Scholar
  76. 76.
    Goehler, al (2005) A protein interaction network links GIT1, an enhancer of huntingtin aggregation, to Huntington’s disease (vol 15, pp. 853, 2004). Mol. Cell19, 287–287CrossRefGoogle Scholar
  77. 77.
    Morsy, al (2008) Charting plant interactomes: possibilities and challenges. Trends Plant Sci.13, 183–191PubMedCrossRefGoogle Scholar
  78. 78.
    Lalonde, S. et al (2008) Molecular and cellular approaches for the detection of protein-protein interactions: latest techniques and current limitations. Plant J.53, 610–635PubMedCrossRefGoogle Scholar
  79. 79.
    Best, al (2007) Localization of protein complexes by pattern recognition. Cell. Electron Microsc.79, 615–638CrossRefGoogle Scholar
  80. 80.
    Adjobo-Hermans, al (2006) Plant G protein heterotrimers require dual lipidation motifs of G alpha and G gamma and do not dissociate upon activation. J. Cell Sci.119, 5087–5097PubMedCrossRefGoogle Scholar
  81. 81.
    Serdyuk, I.N. (2007) Structured proteins and proteins with intrinsic disorder. Mol. Biol.41, 262–277CrossRefGoogle Scholar
  82. 82.
    Dyer, J.M. and Mullen, R.T. (2008) Engineering plant oils as high-value industrial feedstocks for biorefining: the need for underpinning cell biology research. Physiol. Plant.132, 11–22PubMedGoogle Scholar
  83. 83.
    Giegé, P. et al (2003) Enzymes of glycolysis are functionally associated with the mitochondrion in Arabidopsis cells. Plant Cell15, 2140–2151PubMedCrossRefGoogle Scholar
  84. 84.
    Brandina, al (2006) Enolase takes part in a macromolecular complex associated to mitochondria in yeast. Biochim. Biophys. Acta1757, 1217–1228PubMedCrossRefGoogle Scholar
  85. 85.
    Jeffery, C.J. (2005) Mass spectrometry and the search for moonlighting proteins. Mass Spectrom. Rev.24, 772–782PubMedCrossRefGoogle Scholar
  86. 86.
    Moore, B.D. (2004) Bifunctional and moonlighting enzymes: lighting the way to regulatory control. Trends Plant Sci.9, 221–228PubMedCrossRefGoogle Scholar
  87. 87.
    Srere, P.A. (1997) An exception that proves the rule. Trends Biochem. Sci.22, 11–11PubMedCrossRefGoogle Scholar
  88. 88.
    Anderson, L.E. et al (2005) Both chloroplastic and cytosolic phosphofructoaldolase isozymes are present in the pea leaf nucleus. Protoplasma225, 235–242PubMedCrossRefGoogle Scholar
  89. 89.
    Anderson, al (2004) Both chloroplastic and cytosolic phosphoglycerate kinase isozymes are present in the pea leaf nucleus. Protoplasma223, 103–110PubMedGoogle Scholar
  90. 90.
    Anderson, al (2004) Cytosolic glyceraldehyde-3-P dehydrogenase and the B subunit of the chloroplast enzyme are present in the pea leaf nucleus. Protoplasma223, 33–43PubMedCrossRefGoogle Scholar
  91. 91.
    Saslowsky, al (2005) Nuclear localization of flavonoid metabolism in Arabidopsis thaliana<. J. Biol. Chem.280, 23735–23740PubMedCrossRefGoogle Scholar
  92. 92.
    Shimojima, al (2005) Ferredoxin-dependent glutamate synthase moonlights in plant sulfolipid biosynthesis by forming a complex with SQD1. Arch. Biochem. Biophys.436, 206–214PubMedCrossRefGoogle Scholar
  93. 93.
    Matarasso, N. et al (2005) A novel plant cysteine protease has a dual function as a regulator of 1-aminocyclopropane-1-carboxylic acid synthase gene expression. Plant Cell17, 1205–1216PubMedCrossRefGoogle Scholar
  94. 94.
    Pollmann, al (2006) Subcellular localization and tissue specific expression of amidase 1 from Arabidopsis thaliana<. Planta224, 1241–1253PubMedCrossRefGoogle Scholar
  95. 95.
    Lunn, J.E. (2007) Compartmentation in plant metabolism. J. Exp. Bot.58, 35–47PubMedCrossRefGoogle Scholar
  96. 96.
    Jeong, H. et al (2001) Lethality and centrality in protein networks. Nature411, 41–42PubMedCrossRefGoogle Scholar
  97. 97.
    Vélot, C. et al (1997) Model of a quinary structure between Krebs TCA cycle enzymes: a model for the metabolon. Biochemistry (Mosc).36, 14271–14276CrossRefGoogle Scholar

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© Springer Science+Business Media, LLC 2009

Authors and Affiliations

  1. 1.Department of Biological SciencesVirginia TechBlacksburgUSA

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