Catalytic Reactions of the Homogentisate Prenyl Transferase Involved in Plastoquinone-9 Biosynthesis

Abstract

Homogentisate solanesyl transferase (HST) catalyzes the prenylation and decarboxylation of homogentisate to form 2-methyl-6-solanesyl-1,4-benzoquinol, the first intermediate in plastoquinone-9 biosynthesis. In vitro, HST from Spinacia oleracea L., Arabidopsis thaliana, and Chlamydomonas reinhardtii were all found to use not only solanesyl diphosphate but also short chain prenyl diphosphates of 10 -20 carbon atoms as prenyl donors. Surprisingly, with these donors, prenyl transfer was largely decoupled from decarboxylation, and thus the major products were 6-prenyl-1,4-benzoquinol-2-methylcarboxylates rather than the expected 2-methyl-6-prenyl-1,4-benzoquinols. The 6-prenyl-1,4-benzoquinol-2-methylcarboxylates were not substrates for HST-catalyzed decarboxylation, and the enzyme kinetics associated with forming these products appeared quite distinct from those for 2-methyl-6-prenyl-1,4-benzoquinol formation in respect of catalytic rate, substrate K m value, and the pattern of inhibition by haloxydine, a molecule that appeared to act as a dead end mimic of homogentisate. These observations were reconciled into a simple model for the HST mechanism. Here, prenyl diphosphate binds to HST to form at least two alternative complexes that go on to react differently with homogentisate and prenylate it either with or without it first being decarboxylated. It is supposed that solanesyl diphosphate binds tightly and preferentially in the mode that compels prenylation with decarboxylation. Plastoquinone-9 (PQ-9)2 is the major prenylated quinone in chloroplasts. In the thylakoid membrane, it mediates electron flow from photosystem II to the cytochrome b 6 f complex, and the redox state of the PQ-9 pool regulates the expression of a number of nuclear and plastidial genes as well as the activity of some plastidial enzymes (1). Moreover, PQ-9 is required as a cofactor for phytoene desaturation in carotenoid biosynthesis (2). Reflecting its central and unique role in higher plants, a defect in PQ-9 biosynthesis cannot be remedied by other plastidial prenylquinones such as phylloquinone or any of the structurally related intermediates of the vitamin E biosynthetic pathway. Thus the respective mutants of Arabidopsis thaliana display a seedling-lethal phenotype in soil and an albino phenotype on medium supplemented with sucrose (3, 4).PQ-9 comprises an aromatic head group derived from homogentisate and an isoprenoid side chain derived from solanesyl diphosphate (SPP). An HST located in the inner envelope membrane of chloroplasts catalyzes the prenylation and decarboxylation of homogentisate to form 2-methyl-6-solanesyl-1,4-benzoquinol (MSBQ) (Fig. 1) (5), which is then methylated to the reduced form of PQ-9. The HSTs from Chlamydomonas reinhardtii and A. thaliana have been expressed in Escherichia coli and shown to require divalent cations for catalytic activity (6). Other membrane-bound aromatic prenyltransferases exhibit a similar requirement (7). Recent structural studies of the 4-hydroxybenzoate prenyltransferase, UbiA, from E. co...

Cyanobacteria use both p-hydroxybenozate and homogentisate as a precursor of plastoquinone head group

“…PCC 6803. Haloxydine is an inhibitor of HGA prenyltransferases, participating in PQ and a-Toc biosynthesis in higher plants, whereas 4-nitrobenzoate is an inhibitor of HBA prenyltransferase engaged in UQ biosynthesis (Sadre et al 2010;Quinzii et al 2012). Addition of 4-nitrobenzoate significantly slowed down the culture growth, monitored as chlorophyll content in the liquid culture (Fig.…”

Chorismate Pyruvate-Lyase and 4-Hydroxy-3-solanesylbenzoate Decarboxylase Are Required for Plastoquinone Biosynthesis in the Cyanobacterium Synechocystis sp. PCC6803

“…The biosynthesis of PQ in plants occurs in the inner envelope membrane of plastids, where a homogentisate solanesyltransferase catalyzes the decarboxylation and the prenylation of homogentisate (4). The reaction product is subsequently methylated at position 3 so that PQ with a solanesyl (nonaprenyl) side chain is formed (5).…”

AuaA, a Membrane‐Bound Farnesyltransferase from Stigmatella aurantiaca, Catalyzes the Prenylation of 2‐Methyl‐4‐hydroxyquinoline in the Biosynthesis of Aurachins

“…[12,13] The soluble prenyltransferases from fungi and bacteria as well as those involved in the biosynthesis of flavonoids from plants usually use dimethylallyl diphosphate (DMAPP) or geranyl diphosphate (GPP) as prenyl donors, [4,5,7,8] whereas geranylgeranyl diphosphate (GGPP) or diphosphates with longer chains serve as substrates in the biosynthesis of vitamin E, [14] ubiquinones, [9,15] menaquinones, [16] and plastoquinones. [17] Only a few of the known aromatic prenyltransferases accept farnesyl diphosphate (FPP) as a natural prenyl donor. [18,19] It should be mentioned that FPP and GGPP are also substrates for protein prenyltransferases that catalyze the prenylation of the conserved cysteine residue in a CaaX motif of protein and peptide substrates.…”