Plant polysaccharides (cellulose, hemicellulose, pectin, starch) are either direct (i.e. leaf starch) or indirect products of photosynthesis, and they belong to the most abundant organic compounds in nature. Although each of these polymers is made by a specific enzymatic machinery, frequently in different cell locations, details of their synthesis share certain common features. Thus, the production of these polysaccharides is preceded by the formation of nucleotide sugars catalyzed by fully reversible reactions of various enzymes, mostly pyrophosphorylases. These ‘buffering’ enzymes are, generally, quite active and operate close to equilibrium. The nucleotide sugars are then used as substrates for irreversible reactions of various polysaccharide-synthesizing glycosyltransferases (‘engine’ enzymes), e.g. plastidial starch synthases, or plasma membrane-bound cellulose synthase and callose synthase, or ER/Golgi-located variety of glycosyltransferases forming hemicellulose and pectin backbones. Alternatively, the irreversible step might also be provided by a carrier transporting a given immediate precursor across a membrane. Here, we argue that local equilibria, established within metabolic pathways and cycles resulting in polysaccharide production, bring stability to the system via the arrangement of a flexible supply of nucleotide sugars. This metabolic system is itself under control of adenylate kinase and nucleoside-diphosphate kinase, which determine the availability of nucleotides (adenylates, uridylates, guanylates and cytidylates) and Mg2+, the latter serving as a feedback signal from the nucleotide metabolome. Under these conditions, the supply of nucleotide sugars to engine enzymes is stable and constant, and the metabolic process becomes optimized in its load and consumption, making the system steady and self-regulated.

Introduction

Living systems operate far from thermodynamic equilibrium but they maintain the state of homeostasis, which can be reached via optimization of the balance of supply and demand in metabolic pathways [1]. An important role in the supply-demand balance belongs to special thermodynamic ‘buffering’ enzymes that equilibrate fluxes of load and consumption of major metabolites [2–5]. These include ATP synthesis in conjunction with adenylate kinase (AK) reaction [6–8], CO2 fixation buffered by carbonic anhydrase [9] and by equilibration of simple sugars [5], photorespiration [10] and the Krebs cycle [8,11].

The kinetic equilibrium established by ‘buffering’ enzymes is maintained at their high turnover rates, frequently exceeding by several orders of magnitude those of the essential non-equilibrium reactions catalyzed by slow irreversible ‘engine’ enzymes. This arrangement of reversible (buffering) and irreversible (engine) reactions optimizes the balance of supply of substrates and effectors, making the system steady and self-regulated [3,8]. The larger is the contribution of the preceding reversible steps to the optimal operation of slow irreversible enzymes, the closer they are to the equilibrium and the shorter is the transient time for a reversible reaction [12]. The reversible step then becomes rate-determined, and the alteration of the enzyme activity at this step can lead to a corresponding change of the accumulation of product(s) of the irreversible ‘engine’ reaction. Thus, the enzymes that catalyze reactions close to a thermodynamic equilibrium limit the homeostatic flux through the subsequent irreversible step. The theory developed by Heinrich and Rapoport [12] as the basis for the realization of particular metabolic control scenarios, and further elaborated by Fridlyand and Scheibe [13,14], explains the importance of coupling of reversible and irreversible steps in metabolic cycles and pathways for their optimal performance. It represents an important part of the metabolic control theory and provides a possibility of modeling of particular steps of metabolism when the equilibrium constants and concentrations of metabolites, in a given compartment, for individual reactions are determined.

Recently [8], we discussed how the equilibria of coenzyme nucleotides and substrates and their feedbacks generate computable rules that govern the plant metabolome and provide optimal conditions for the stable non-equilibrium fluxes of the processes of ATP synthesis, CO2 fixation and mitochondrial respiration. In the current paper, we expand this discussion to the analysis of how the equilibria in enzymatic reactions, in particular, established with the participation of adenylate, uridylate and guanylate nucleotides, govern the synthesis of various polysaccharides through the supply of nucleotide sugars. To our knowledge, the paper is the first attempt to describe polysaccharide synthesis in plants from the point of view of thermodynamic equilibria and buffering. This opinion paper outlines the current knowledge of the de novo formation of nucleotide sugars and defines the applicability of the thermodynamic buffering concept for understanding the biochemical mechanism and regulation of polysaccharide synthesis. This includes the following key points: (i) reversibility of nucleotide sugar synthesizing (‘buffering’) reactions, (ii) irreversibility of polysaccharide synthases (‘engine’ reactions) and (iii) the balance of reversible and irreversible steps in the context of regulatory mechanisms imposed by AK and nucleoside-diphosphate kinase (NDPK) to control the availability of various nucleotides and free magnesium as a feedback signal. The maintenance of polysaccharide synthesis by local concentrations of nucleoside triphosphates (NTPs), pyrophosphate (PPi) and Mg2+, under conditions of the stable non-equilibrium flux of newly produced nucleotide sugars toward irreversible reactions catalyzed by glycosyltransferases, will be discussed.

Plant polysaccharides and their metabolism

Plant polysaccharides are the most abundant biomaterials in nature [15]. The most prominent among them — cellulose, hemicellulose and pectin — are located in cell walls and are essential for cell wall architecture and properties. On the other hand, starch, another highly abundant polysaccharide, is localized in plastids and is used as an energy storage material and source of carbon skeletons for metabolism. Although they differ in enzymatic details of their synthesis and are frequently synthesized in different cell compartments, the general principles involved in polysaccharide formation are similar.

In leaves (source tissue), the formation of polysaccharides is preceded by a series of reactions from triose-P, the simplest of sugars formed by the Calvin–Benson cycle, to specific nucleotide sugars, which serve as direct oligo- and polysaccharide precursors. Thus, UDP-d-glucose (UDP-Glc) is produced in the cytosol for the synthesis of sucrose (Suc) and cell wall polysaccharides, whereas ADP-Glc is synthesized in chloroplasts for starch production [16,17]. Suc is predominantly synthesized by coupled reactions of Suc-P synthase (SPS) and Suc-P phosphatase (SPPase), where SPS carries out a reversible formation of Suc-6-P and UDP from UDP-Glc and Fru-6-P, whereas the phosphatase is irreversible [17,18] (Figure 1A). Both SPS and the phosphatase interact physically with each other, which impacts soluble carbohydrate pools and affects carbon partitioning to starch [19]. In this way, metabolic channeling from UDP-Glc to Suc in the cytosol is ensured. UDP-Glc is formed by UDP-Glc pyrophosphorylase (UGPase) from Glc-1-P and UTP, with Glc-1-P derived from fructose-6-P (Fru-6-P) via cytosolic activities of phosphoglucoisomerase (PGI) and phosphoglucomutase (PGM). Since all these enzymes are freely reversible, one could assume that there is an equilibrium set from Fru-6-P all the way to Suc-6-P (Figure 1A) [20].

Primary metabolism in source (leaves) (A) and sink (non-photosynthetic) (B) tissues.

Figure 1.
Primary metabolism in source (leaves) (A) and sink (non-photosynthetic) (B) tissues.

Blue lines refer to freely reversible reactions, red lines — irreversible reactions. Arrows point toward the actual carbon flow driven by irreversible reactions. Dashed lines show the involvement of two or more reactions. 1 — CesA, CalS and hemicellulose synthases; 2 — pectin synthases; 3 — starch synthase isozymes and branching/debranching enzyme. Abbreviations: AGP, ADP-Glc pyrophosphorylase; F6P, Fru-6-P; FrK, fructokinase; G1P, Glc-1-P; G6P, Glc-6-P; HXK, hexokinase; Inv, invertase; PGI, phosphoglucoisomerase; PGM, phosphoglucomutase; SPase, Suc-P phosphatase; SPS, Suc-P synthase; SuSy, sucrose synthase; UGP, UDP-Glc pyrophosphorylase.

Figure 1.
Primary metabolism in source (leaves) (A) and sink (non-photosynthetic) (B) tissues.

Blue lines refer to freely reversible reactions, red lines — irreversible reactions. Arrows point toward the actual carbon flow driven by irreversible reactions. Dashed lines show the involvement of two or more reactions. 1 — CesA, CalS and hemicellulose synthases; 2 — pectin synthases; 3 — starch synthase isozymes and branching/debranching enzyme. Abbreviations: AGP, ADP-Glc pyrophosphorylase; F6P, Fru-6-P; FrK, fructokinase; G1P, Glc-1-P; G6P, Glc-6-P; HXK, hexokinase; Inv, invertase; PGI, phosphoglucoisomerase; PGM, phosphoglucomutase; SPase, Suc-P phosphatase; SPS, Suc-P synthase; SuSy, sucrose synthase; UGP, UDP-Glc pyrophosphorylase.

In non-photosynthetic (sink) tissues, polysaccharide synthesis is linked with Suc metabolism via sucrose synthase (SuSy) and invertase (Inv) [21] (Figure 1B). Whether it is SuSy or Inv which takes the major role in Suc breakdown depends on plant species, tissue type, abiotic conditions, etc. [22]. For instance, SuSy role in cell wall biosynthesis was shown in cotton [23] and bean [24], whereas studies on genetically modified plants demonstrated that cytosolic Inv plays central role in cellulose biosynthesis and carbon allocation in Arabidopsis [25] and hybrid aspen [26]. In case Inv is prevalent in the breakdown of Suc in the sink, then the equilibrium situation applies from Glc-6-P to ADP-Glc (starch formation) or from Glc-6-P (or Fru-6-P) all the way to UDP-Glc (cell wall polysaccharides synthesis) (Figure 1B).

Cell wall polysaccharides

Cellulose forms long unbranched chains of 7000–15 000 Glc units, linked via a β1,4 bond. Hemicellulose, in contrast, is a heteropolymer with a lot of side chains and is composed of pentoses, mostly d-xylose (Xyl), as well as hexoses, such as Glc, d-mannose (Man), d-galactose (Gal) and others. Side sugars may include d-glucuronic acid (GlcA), l-arabinose (Ara), l-fucose (Fuc) and other sugars. According to the main sugar residue in its backbone, hemicelluloses are classified as xylans, glucans, mannans and galactans. Hemicellulose polymers are shorter than those of cellulose and usually consist of 50–3000 sugar units. Pectin, another component of plant cell walls, is generally a polymer of d-galacturonic acid (GalA), with a variety of sugar side-chains attached. Those branches may contain Ara, Gal, Fuc and other sugar residues. Cell walls contain various types of hemicelluloses and pectins, depending on plant species, developmental stage, organ/tissue type, etc. [27–29].

The synthesis of cellulose occurs at the plasmalemma and is carried out by a complex of proteins called cellulose synthase (CesA). In the reaction, CesA uses UDP-Glc as a glucosyl donor to a [(1 → 4)-β-d-glucosyl]n acceptor, extending the cellulose chain by one Glc at a time and producing one UDP per reaction [24]. Plasmalemma is also the site of synthesis of callose, a polymer also composed entirely of Glc molecules, linked by β1,3-linkages and with some β1,6-branches, and produced by callose synthase (CalS) [30]. In the sink, the UDP-Glc used by both CesA and CalS comes from SuSy-catalyzed breakdown of Suc [23,24,31], but UGPase could also contribute, at least theoretically (Figure 1B) [25,32]. SuSy was found to bind directly to the catalytic site of CesA complex in bean plasmalemma fraction [24].

In contrast with cellulose and callose, the synthesis of hemicellulose and pectin occurs in ER/Golgi, and then the polysaccharides are transported in vesicles to be unloaded to the cell walls [33].

Starch

Starch is yet another carbohydrate polymer composed entirely of Glc units, but connected via α1,4-link (backbone chain) and α1,6-link (for the branching points). Although not as abundant as cell wall polysaccharides, it is nevertheless extremely important from the energy point of view, as it can be synthesized and stored and then easily broken down to Glc units. The Glc then can be used both as a fuel to produce energy during respiration, but also as the provider of carbon skeletons for the whole of plant metabolism. In contrast with cell wall polysaccharides, starch is always formed in plastids, in both source and sink tissues. In the leaves, starch is made during light conditions, and is broken down (to maltose) in the darkness. Recently, there has been evidence that maltose can also be produced in the light, pointing out to both synthesis and mobilization of starch in the leaves during photosynthesis [34,35]. In the sink, starch can be synthesized or broken down regardless of light/dark conditions, but depending on the growth/developmental stage [36,37]. Similar to cell wall polysaccharides, the immediate enzymatic machinery for starch synthesis is the same in both source and sink tissues, but there are differences in the origin of carbon. In leaves, it is predominantly triose-P originating from the Calvin–Benson cycle, whereas in non-photosynthetic tissues it is mainly Suc transported from leaves in the phloem [36,37] (Figure 1).

The de novo formation of nucleotide sugars is carried out by reversible reactions

Table 1 presents a list of enzymes de novo producing NDP-sugars, which are precursors for polysaccharide formation. The list includes several pyrophosphorylases, all using NTP and various sugar-1-phosphates as substrates, as well as SuSy and two nucleotide sugar phosphorylases: ADP-Glc phosphorylase (AGase) and GDP-Gal phosphorylase (VTC2/VTC5), all using NDP as one of the substrates. It also contains CMP-3-deoxy-d-manno-octulosonate synthetase (CKS), producing a unique NMP-linked sugar derivative CMP-keto-deoxyoctulosonate (CMP-Kdo) [38]. All these reactions are reversible, some are quite active (e.g. SuSy, UGPase, AGPase) and most of them are localized in the cytosol.

Table 1
De novo nucleotide sugar synthesis, major polysaccharide synthases and equilibrium constants of relevant enzymes
NDP-sugar formation and other relevant reactionsKeqMajor use in polysaccharide synthesisReferences
UDP-sugar synthesis
 UGPase UTP + Glc-1-P ↔ UDP-Glc + PPi 0.3–0.4 Cellulose, callose, hemicellulose, PNG Meng et al. [51]; Decker et al. [121
 USPase UTP + Sugar-1-P ↔ UDP-Sugar + PPi 0.2–0.5 Pectin, hemicellulose, PNG, POG Kotake et al. [41
 UAGPase UTP + GlcNAc-1-P ↔ UDP-GlcNAc + PPi 0.15–0.3 PNG, POG Mengin-Lecreulx and van Heijenoort [122
 SuSy UDP + Sucrose ↔ UDP-Glc + Fru 0.2–0.6 Cellulose, callose, hemicellulose, PNG Geigenberger and Stitt [123
ADP-Glc synthesis
 AGPase ATP + Glc-1-P ↔ ADP-Glc + PPi 0.5 Starch Preiss [16]; Kleczkowski [60
 SuSy ADP + Sucrose ↔ ADP-Glc + Fru 0.2–0.61 Starch Geigenberger and Stitt [123
 AGase ADP + Glc-1-P ↔ ADP-Glc + Pi 0.025–0.2 Starch McCoy et al. [75
GDP-sugar synthesis
 GMPase GTP + Man-1-P ↔ GDP-Man + PPi 0.4 Hemicellulose, PNG, POG Szumilo et al.[124
 FKGP GTP + Fuc-1-P ↔ GDP-Fuc + PPi Reversible2 Pectin, hemicellulose, PNG, POG Kotake et al. [125
 VTC2/5 GDP + Glc-1-P ↔ GDP-Glc + Pi Reversible2 Hemicellulose Linster et al. [126
CMP-Kdo synthesis
 CKS CTP + Kdo ↔ CMP-Kdo + PPi Reversible2 Pectin Ghalambor and Heath [82
Polysaccharide synthases 
 CesA UDP-Glc + (Glc)n → (Glc)n+1 + UDP Irreversible Cellulose Nixon et al. [83
 CalS UDP-Glc + (Glc)n → (Glc)n+1 + UDP Irreversible Callose Cai et al. [31
 HS NDP-Sugar + (Sugar)n → (Sugar)n+1 + NDP Irreversible Hemicellulose Scheller and Ulvskov [29
 PS NDP-Sugar + (Sugar)n → (Sugar)n+1 + NDP Irreversible Pectin Harholt et al. [28
 StSy ADP-Glc + (Glc)n → (Glc)n+1 + ADP Irreversible Starch Preiss [16
Other key reactions 
 AK ADP + MgADP ↔ MgATP + AMP 6.0 Indirect (all polysaccharides) Kleczkowski and Randall [92
 NDPK ATP + NDP ↔ NTP + ADP 1.0 Indirect (all polysaccharides) Moffatt and Ashihara [127
 PGM Glc-6-P ↔ Glc-1-P 0.05 Indirect (all polysaccharides) Mutuku and Nose [128
 PGI Glc-6-P ↔ Fru-6-P 0.4–0.5 Indirect (all polysaccharides) Mutuku and Nose [128
 SPS UDP-Glc + Fru-6-P ↔ Suc-6-P + UDP 5–62 Indirect (all polysaccharides) Lunn and ap Rees [18
 SPPase Suc-6-P → Suc + Pi Irreversible Indirect (all polysaccharides) Taiz et al. [36
 Inv Suc → Glc + Fru Irreversible Indirect (all polysaccharides) Taiz et al. [36
 HK Glc + ATP → Glc-6-P + ADP Irreversible Indirect (all polysaccharides) Taiz et al. [36
 FK Fru + ATP → Fru-6-P + ADP Irreversible Indirect (all polysaccharides) Taiz et al. [36
NDP-sugar formation and other relevant reactionsKeqMajor use in polysaccharide synthesisReferences
UDP-sugar synthesis
 UGPase UTP + Glc-1-P ↔ UDP-Glc + PPi 0.3–0.4 Cellulose, callose, hemicellulose, PNG Meng et al. [51]; Decker et al. [121
 USPase UTP + Sugar-1-P ↔ UDP-Sugar + PPi 0.2–0.5 Pectin, hemicellulose, PNG, POG Kotake et al. [41
 UAGPase UTP + GlcNAc-1-P ↔ UDP-GlcNAc + PPi 0.15–0.3 PNG, POG Mengin-Lecreulx and van Heijenoort [122
 SuSy UDP + Sucrose ↔ UDP-Glc + Fru 0.2–0.6 Cellulose, callose, hemicellulose, PNG Geigenberger and Stitt [123
ADP-Glc synthesis
 AGPase ATP + Glc-1-P ↔ ADP-Glc + PPi 0.5 Starch Preiss [16]; Kleczkowski [60
 SuSy ADP + Sucrose ↔ ADP-Glc + Fru 0.2–0.61 Starch Geigenberger and Stitt [123
 AGase ADP + Glc-1-P ↔ ADP-Glc + Pi 0.025–0.2 Starch McCoy et al. [75
GDP-sugar synthesis
 GMPase GTP + Man-1-P ↔ GDP-Man + PPi 0.4 Hemicellulose, PNG, POG Szumilo et al.[124
 FKGP GTP + Fuc-1-P ↔ GDP-Fuc + PPi Reversible2 Pectin, hemicellulose, PNG, POG Kotake et al. [125
 VTC2/5 GDP + Glc-1-P ↔ GDP-Glc + Pi Reversible2 Hemicellulose Linster et al. [126
CMP-Kdo synthesis
 CKS CTP + Kdo ↔ CMP-Kdo + PPi Reversible2 Pectin Ghalambor and Heath [82
Polysaccharide synthases 
 CesA UDP-Glc + (Glc)n → (Glc)n+1 + UDP Irreversible Cellulose Nixon et al. [83
 CalS UDP-Glc + (Glc)n → (Glc)n+1 + UDP Irreversible Callose Cai et al. [31
 HS NDP-Sugar + (Sugar)n → (Sugar)n+1 + NDP Irreversible Hemicellulose Scheller and Ulvskov [29
 PS NDP-Sugar + (Sugar)n → (Sugar)n+1 + NDP Irreversible Pectin Harholt et al. [28
 StSy ADP-Glc + (Glc)n → (Glc)n+1 + ADP Irreversible Starch Preiss [16
Other key reactions 
 AK ADP + MgADP ↔ MgATP + AMP 6.0 Indirect (all polysaccharides) Kleczkowski and Randall [92
 NDPK ATP + NDP ↔ NTP + ADP 1.0 Indirect (all polysaccharides) Moffatt and Ashihara [127
 PGM Glc-6-P ↔ Glc-1-P 0.05 Indirect (all polysaccharides) Mutuku and Nose [128
 PGI Glc-6-P ↔ Fru-6-P 0.4–0.5 Indirect (all polysaccharides) Mutuku and Nose [128
 SPS UDP-Glc + Fru-6-P ↔ Suc-6-P + UDP 5–62 Indirect (all polysaccharides) Lunn and ap Rees [18
 SPPase Suc-6-P → Suc + Pi Irreversible Indirect (all polysaccharides) Taiz et al. [36
 Inv Suc → Glc + Fru Irreversible Indirect (all polysaccharides) Taiz et al. [36
 HK Glc + ATP → Glc-6-P + ADP Irreversible Indirect (all polysaccharides) Taiz et al. [36
 FK Fru + ATP → Fru-6-P + ADP Irreversible Indirect (all polysaccharides) Taiz et al. [36

AGase, ADP-Glc phosphorylase; AGPase, ADP-Glc pyrophosphorylase; AK, adenylate kinase; CalS, callose synthase; CesA, cellulose synthase; CKS, CMP-Kdo synthetase; FK, fructose kinase; FKGP, Fuc kinase:GDP-Fuc pyrophosphorylase; GMPase, GDP-Man pyrophosphorylase (VTC1); HK, hexokinase; HS, hemicellulose synthases (generally); Inv, invertase; NDPK, nucleoside-diphosphate kinase; PGI, phosphoglucoisomerase; PGM, phosphoglucomutase; PNG, protein N-glycosylation; POG, protein O-glycosylation; PS, pectin synthases (generally); SPS, sucrose-P synthase; SPPase, sucrose-P phosphatase; StSy, starch synthase isozymes; SuSy, sucrose synthase; UAGPase, UDP-GlcNAc (UDP-GalNAc) pyrophosphorylase; UGPase, UDP-Glc pyrophosphorylase; USPase, UDP-sugar pyrophosphorylase; VTC2 and VTC5, GDP-l-Gal (GDP-d-Glc) phosphorylase.

1

Assuming the same Keq as with UDP-Glc as one of the reactants;

2

Keq unknown, but the reaction reported as reversible.

The importance of reversibility is especially evident for SuSy, which in source tissues is believed to be involved in Suc formation, although to a lesser extent than SPS, and in the sink tissues in the breakdown of Suc to UDP-Glc (and/or ADP-Glc) and Fru to make the carbon available for synthesis reactions, e.g. of polysaccharides, and for respiration [17,32] (Figure 1). Similarly distinct are the roles of UGPase, which in source tissues makes UDP-Glc for Suc and cell wall synthesis, whereas in the sink it likely uses UDP-Glc, the product of SuSy, to make UTP and Glc-1-P to replenish energy and carbon skeletons [32,39] (Figure 1). Such a ‘coupling’ between SuSy and UGPase in the sink makes metabolism more energy efficient under conditions of limiting oxygen concentration (hypoxia), as in non-photosynthetic tissues [4].

UDP-sugar metabolism

Among nucleotide sugars produced by the reactions shown in Table 1, UDP-Glc is by far the most abundant [40], being predominantly synthesized by UGPase and SuSy, but also by non-specific reactions of UDP-sugar pyrophosphorylase (USPase) and UDP-GlcNAc (UDP-GalNAc) pyrophosphorylase (UAGPase) [32,41–45]. Whereas UGPase appears more or less specifically involved in UDP-Glc formation, USPase is a highly promiscuous enzyme and, in addition to UDP-Glc, it can also produce UDP-Gal, UDP-GlcA, UDP-GalA, UDP-Ara, UDP-Xyl and UDP-α-d-Fuc [41,42,44,46–48], using a given sugar-1-P and UTP as substrates. UAGPase, on the other hand, is involved in UDP-GlcNAc and UDP-GalNAc biosynthesis, which are important precursors for N- and O-linked protein glycosylation [43,49,50].

Plants contain two isozymes of UGPase, coded by distinct genes, both isozymes sharing similar properties [51]. In transgenic potato plants, ‘antisense’ inhibition of tuber UGPase down to 4% of its rate in wild type plants had no effect on Suc and starch levels, strongly suggesting that the enzyme is not rate-limiting in the tubers [52]. Studies with mutants of Arabidopsis with knockouts and knockdowns of UGPase activity brought about similar results; i.e. a decrease in UGPase activity down to 15% had no apparent effects on soluble sugar and starch content, and there were no differences in cell wall composition/content between the wild type and the mutants [53]. Only a full double UGPase mutant, with 6% of UGPase-like activity left, was severely affected, and could only be sustained when grown on 1.5% Suc and 1.5% UDP-Glc [54]. This strongly indicates that UGPase is not a rate-limiting step in metabolism, but that it is essential for normal growth/development. The UGPase-like activity in the double knockout most likely can be attributed to USPase and UAGPase, which use Glc-1-P as one of their non-specific substrates [44] and to a unique chloroplastic UGPase, which is involved in sulfolipid biosynthesis [55], and belongs to a separate family of proteins, distinct from cytosolic UGPases [56]. The exact in planta contribution of each of these enzymes to the formation of UDP-Glc is difficult to determine, but it could be clarified using specific inhibitors in assays in crude plant extracts [57].

Once UDP-Glc is produced, either by UGPase or SuSy, there are several nucleotide sugar inter-conversion enzymes, which utilize it to synthesize UDP-Gal, UDP-GlcA and other UDP-sugars [44,45,50,58]. Several of these inter-conversion enzymes produce the same nucleotide sugars as USPase, underlying the complexity and, frequently, the redundancy of nucleotide sugar metabolism [45].

In addition to its metabolic function, UDP-Glc (and perhaps other UDP-sugars) may have signaling roles, acting as an extracellular signal that is perceived as a damage-associated molecular pattern [59].

ADP-Glc metabolism and starch synthesis

Starch synthesis is under strong control of ADP-Glc pyrophosphorylase (AGPase), the first committed step of starch synthesis. AGPase is activated by 3-phosphoglycerate (PGA), the first stable product of the Calvin–Benson cycle, and inhibited by inorganic phosphate (Pi), which is a crucial substrate for photophosphorylation and oxidative (respiratory) phosphorylation [16,60]. The PGA/Pi ratio was demonstrated to regulate leaf AGPase activity in the same way as it regulates changes in the amount of starch in chloroplasts [61], strongly suggesting that AGPase constitutes a rate-limiting step, which directly controls starch formation in leaves. Based on this and other results, it has been calculated that steady-state levels of the Calvin–Benson cycle intermediates in the chloroplasts are controlled by the external level of Pi and that the control of starch synthesis by PGA/Pi ratio via AGPase makes the photosynthetic system more stable in a wide range of external Pi concentrations [62].

In addition to the classical starch synthesis pathway in leaves, where ADP-Glc originates from triose-P produced by the Calvin–Benson cycle (Figure 1A) [16], plants possess also additional/alternative pathway(s) of transitory starch biosynthesis, wherein (i) ADP-Glc produced in the cytosol enters the chloroplast for its subsequent conversion into starch [63] and (ii) starch synthesis and breakdown occur simultaneously during illumination [34]. In the latter case, plastidial PGM and AGPase (but not plastidial PGI) play an important role in recycling back to starch its own breakdown products [63].

Besides AGPase, none of the other NDP-sugar producing reactions appear to be under metabolic regulation, aside from product inhibition of some pyrophosphorylases [39,64,65]. Other types of regulation, although none of them as prominent as PGA/Pi regulation of AGPase, may involve protein phosphorylation (AGPase, UGPase, SuSy), and redox regulation (AGPase, UGPase), as well as protein oligomerization (UGPase). Details on those, as well as on the regulation of expression of corresponding genes, have been reviewed elsewhere [32,45,50,58].

Other than AGPase reaction, ADP-Glc can be also produced by SuSy, when it uses ADP instead of UDP for Suc breakdown. The production of ADP-Glc by SuSy is well documented [66,67]. On the other hand, the exact in vivo extent of production of ADP-Glc versus UDP-Glc by SuSy is unknown, but it could be substantial [68,69].

In cereal seed endosperm, which contains a highly active cytosolic isozyme of AGPase [70,71], its activity is most likely directly linked to that of UGPase which, in turn, can be connected to SuSy activity, providing a direct connection between Suc breakdown and starch synthesis in this tissue [69–73]. In such a scheme, three enzymes — SuSy, UGPase and AGPase — are metabolically linked and equilibrated from Suc to ADP-Glc (Figure 2). The latter is then imported to the plastids by BT1 transporter, in exchange with ADP, and used by starch synthase for amylose synthesis [74]. On the other hand, if SuSy produces ADP-Glc instead of UDP-Glc, SuSy will feed ADP-Glc directly to BT1 translocator for starch synthesis in the plastids (Figure 2). Thus, regardless of whether SuSy produces UDP-Glc or ADP-Glc, Suc breakdown in cereal seed endosperm serves mainly toward starch synthesis, and the whole conversion of Suc to ADP-Glc can be considered as being under equilibrium. Also, when considering the whole carbon flow from the SuSy-broken Suc to starch, there is no net energy expenditure. This is true, regardless of whether ADP-Glc is formed by cytosolic SuSy (1 UDP used and 1 ADP produced) or by cytosolic AGPase (1 ATP used and 1 ADP produced per 1 UTP produced and 1 UDP used) [71–73] (Figure 2). In the leaves, however, starch production derived from the Calvin–Benson cycle activity is an energy-requiring process (1 ATP spent for the production of ADP-Glc by chloroplastic AGPase) [16].

Metabolic connection between Suc breakdown by SuSy and starch synthesis in cereal seed endosperm.

Figure 2.
Metabolic connection between Suc breakdown by SuSy and starch synthesis in cereal seed endosperm.

Blue lines refer to freely reversible reactions, red lines — irreversible reactions. Arrows point toward the actual carbon flow driven by irreversible reactions. Please note that SuSy reaction is presented as it can produce both UDP-Glc and ADP-Glc. For clarity, the scheme contains no details of amyloplastic isozyme of AGPase (much less active than the cytosolic isozyme in seed endosperm) [71]. Abbreviations: ADPG, ADP-Glc; AGP, ADP-Glc pyrophosphorylase; Bt1, adenylate translocator; G1P, Glc-1-P; PPi, pyrophosphate; StSy, starch synthase isozymes; SuSy, sucrose synthase; UDPG, UDP-Glc; UGP, UDP-Glc pyrophosphorylase.

Figure 2.
Metabolic connection between Suc breakdown by SuSy and starch synthesis in cereal seed endosperm.

Blue lines refer to freely reversible reactions, red lines — irreversible reactions. Arrows point toward the actual carbon flow driven by irreversible reactions. Please note that SuSy reaction is presented as it can produce both UDP-Glc and ADP-Glc. For clarity, the scheme contains no details of amyloplastic isozyme of AGPase (much less active than the cytosolic isozyme in seed endosperm) [71]. Abbreviations: ADPG, ADP-Glc; AGP, ADP-Glc pyrophosphorylase; Bt1, adenylate translocator; G1P, Glc-1-P; PPi, pyrophosphate; StSy, starch synthase isozymes; SuSy, sucrose synthase; UDPG, UDP-Glc; UGP, UDP-Glc pyrophosphorylase.

ADP-Glc can also be produced from ADP and Glc-1-P via a scarcely studied AGase, a phosphorylase, which is widely distributed in plant tissues and carries a reversible reaction, which nevertheless at some conditions favors the utilization of ADP-Glc [75] (Table 1). Thus, the enzyme may be involved in the breakdown of ADP-Glc, in contrast with AGPase. In this way, the AGase role may be analogous to ADP-Glc pyrophosphatase [76,77], belonging to a nucleotide pyrophosphatase/phosphodiesterase family of proteins, which under certain conditions breaks down ADP-Glc, and diverts the flow of carbon from starch synthesis [78].

GDP-sugars and other nucleotide sugars

Besides ADP-Glc and UDP-sugars, plants contain also several GDP-sugars formed by specific enzymes. Two of these are pyrophosphorylases, using GTP as nucleotide and producing GDP-Man and GDP-Fuc, respectively, that are utilized in pectin and hemicellulose formation, and one is a phosphorylase, using GDP as one of the substrates and producing GDP-Glc for hemicellulose synthesis (Table 1). All these reactions are reversible.

Although CDP-Glc pyrophosphorylase-like activity has been reported in plant tissues [79], the roles of plant CDP-sugars remain unknown and largely unexplored. This is in contrast with bacteria, where CDP-sugars are involved in lipopolysaccharide production [80]. However, plants contain CMP-Kdo, an unusual NMP-linked sugar involved in pectin rhamnogalacturonan II formation, made by CKS (Table 1), an ancient enzyme thought to be acquired by plants from Gram-negative eubacteria via horizontal gene transfer [38,50,81]. Similar to pyrophosphorylases, the enzyme carries out a reversible reaction, although the equilibrium of the reaction lies in favor of CMP-Kdo formation [82].

Polysaccharide synthases are slow and abundant enzymes carrying out irreversible reactions

The availability of nucleotide sugars and the activities of specific glycosyltransferases that use them for polysaccharide production are the key factors controlling the amounts of cell wall carbohydrates and starch. Compared with their substrate concentrations, the glycosyltransferases (synthases) are frequently present at relatively high amounts. For instance, a characteristic ‘rosette’-like structure of cellulose synthase can be seen on electron microscopy photographs of the isolated plasma membrane [83], whereas granule-bound starch synthase is the most abundant protein found in crude starch preparations [84]. Although this is speculation only, the presence of high amount of a given enzyme suggests that its reaction is slow per enzyme molecule. An analogy can be drawn with Rubisco, the most abundant enzyme/protein in nature, carrying out CO2 fixation during the Calvin–Benson cycle in chloroplasts, where the apparent reason for its abundance is its very slow catalytic rate [85]. The slowness of abundant enzymes may be also related to the evolutionary tradeoff for carrying out a very specific reaction [86], e.g. given the multitude of distinct Glc-only containing polymers differing only in the links between Glc molecules (β1–4 in cellulose chain; β1–3 in callose; α1–4 in starch, etc.). Both CesA and CalS, as well as enzymes synthesizing different types of hemicelluloses, are grouped into a single GT2 family [87], presumably arising from a common ancient precursor.

Aside from CesA and CalS, which are localized at the plasmalemma [31], most of the polysaccharide synthesis machinery involved in cell wall formation is localized in ER/Golgi. Thus, the enzymes involved are physically separated by a membrane from the cytosol where most of the nucleotide sugars are made. Those nucleotide sugars need to be transported to ER/Golgi on specific translocators. Up till now, several ER/Golgi nucleotide sugar transporters have been identified in Arabidopsis, based on experimental studies and on comparisons to phylogenetic trees with known transporters from other organisms [88]. Some of the transporters are more or less specific for a given nucleotide sugar, e.g. GDP-l-Fuc transporter [89], whereas others can transport two or more types of these compounds [88]. For a given compound to be transported, as the translocators work usually only unidirectionally, the transport step may be regarded as an irreversible reaction that effectively removes a compound from a given metabolite pool [90].

A similar role can be envisioned for the plastidic BT1 transporter and starch synthesis. The BT1 transporter is the most abundant protein in amyloplast membranes [91] and, from a metabolic/enzymatic point of view, its involvement in the import of ADP-Glc can be considered as an irreversible reaction, driving the flow of carbon from Suc in the cytosol into plastids (Figure 2).

Polysaccharide synthesis via thermodynamic buffering

In the thermodynamic buffering concept [2], a prominent role is played by AK which carries out the following reaction: MgADP + ADP ↔ MgATP + AMP. The enzyme maintains the equilibrium of Mg-bound and free adenylates as well as that of Mg2+ in chloroplasts, cytosol and in the intermembrane space in mitochondria [92]. This, in turn, has profound effects on the operation of major metabolic pathways and their regulation [4,7,93,94]. While ATP is the only NTP synthesized during photophosphorylation and oxidative (respiratory) phosphorylation [95], the AK-mediated equilibrium of adenylates is linked to equilibria of guanylates, uridylates and cytidylates via sets of specific kinases that transform a part of stored energy to other nucleoside triphosphates — GTP, UTP and CTP. Such enzymes as guanylate kinase, uridylate kinase (UMP kinase) and CMP kinase, operating together with NDPK, are important players providing the balance of nucleotides for general metabolism [8,96–98]. The role of NDPK seems especially important since, similar to AK, it is a ‘buffering’ enzyme, but contrary to plant AK [99] it can react with all nucleotides, although with different efficiency [8,98]. The reaction of NDPK can be presented as MgATP + NDP ↔ ADP + MgNTP. Both AK and NDPK are functionally coupled to the oxidative phosphorylation system [100] and they exhibit metabolic cooperation [101]. Equilibrium reactions of AK and NDPK, in the context of polysaccharide formation via thermodynamic buffering, are shown in Figure 3.

Equilibrium reactions of the nucleotide metabolome optimizing the synthesis of polysaccharides in plants.

Figure 3.
Equilibrium reactions of the nucleotide metabolome optimizing the synthesis of polysaccharides in plants.

The reactions involved: 1 — ATP synthase; 2 — adenylate kinase (AK); 3 — nucleoside-diphosphate kinase (NDPK); 4 — guanylate kinase; 5 — UMP kinase; 6 — CMP kinase and NDPK; 7 — ADP-Glc pyrophosphorylase (AGPase); 8 — ADP-Glc phosphorylase (AGase); 9 — UDP-Glc (or other UDP-sugar) pyrophosphorylase; 10 — GDP-fucose or GDP-mannose pyrophosphorylase (FKGP or GMPase); 11 — inorganic pyrophosphatase (PPase) and vacuolar H+-PPase; 12 — GDP-Glc phosphorylase (VTC2/5); 13 — CMP-Kdo synthetase (CKS). 14 — starch synthase isozymes and branching/debranching enzyme; 15 — cellulose synthase (CesA); 16 — callose synthase (CalS); 17, 18, 20, 22, 23 — hemicellulose synthases (generally); 19, 21, 24 — pectin synthases (generally). Dashed lines show the supply of ADP for ATP synthesis. Nucleoside monophosphates are shown in purple, Mg2+ equilibrated by AK and participating in most reactions is in green. Reversible reactions are in blue, irreversible — in red. SugX represents all sugars other than Glc that can be de novo converted to UDP-sugar. The figure does not show reactions of SuSy, which produces UDP-Glc (or ADP-Glc) and fructose from UDP (or ADP) and sucrose, nor the reactions of UAGPase which produces UDP-GlcNAc (or UDP-GalNAc) and PPi from UTP and GlcNAc-1-P (or GalNAc-1-P).

Figure 3.
Equilibrium reactions of the nucleotide metabolome optimizing the synthesis of polysaccharides in plants.

The reactions involved: 1 — ATP synthase; 2 — adenylate kinase (AK); 3 — nucleoside-diphosphate kinase (NDPK); 4 — guanylate kinase; 5 — UMP kinase; 6 — CMP kinase and NDPK; 7 — ADP-Glc pyrophosphorylase (AGPase); 8 — ADP-Glc phosphorylase (AGase); 9 — UDP-Glc (or other UDP-sugar) pyrophosphorylase; 10 — GDP-fucose or GDP-mannose pyrophosphorylase (FKGP or GMPase); 11 — inorganic pyrophosphatase (PPase) and vacuolar H+-PPase; 12 — GDP-Glc phosphorylase (VTC2/5); 13 — CMP-Kdo synthetase (CKS). 14 — starch synthase isozymes and branching/debranching enzyme; 15 — cellulose synthase (CesA); 16 — callose synthase (CalS); 17, 18, 20, 22, 23 — hemicellulose synthases (generally); 19, 21, 24 — pectin synthases (generally). Dashed lines show the supply of ADP for ATP synthesis. Nucleoside monophosphates are shown in purple, Mg2+ equilibrated by AK and participating in most reactions is in green. Reversible reactions are in blue, irreversible — in red. SugX represents all sugars other than Glc that can be de novo converted to UDP-sugar. The figure does not show reactions of SuSy, which produces UDP-Glc (or ADP-Glc) and fructose from UDP (or ADP) and sucrose, nor the reactions of UAGPase which produces UDP-GlcNAc (or UDP-GalNAc) and PPi from UTP and GlcNAc-1-P (or GalNAc-1-P).

The formation of different NTPs not only generates the substrates for RNA (and DNA) synthesis, but also makes possible an independent regulation of the supply of energy for distinct metabolic pathways that are involved in protein and DNA (mostly GTP is required), polysaccharide (UTP and ATP) and lipid (CTP) synthesis [102]. This provides an additional control in the cell over processes fulfilling useful work [103]. In particular, the equilibria in the reactions involving adenylates, uridylates and guanylates provide sufficient concentrations of substrates for the irreversible reactions of polysaccharide synthesis. It seems important to emphasize that Km values for substrates of NDP-sugar producing enzymes, when considering both directions of the reaction, are generally lower than the actual concentrations of these compounds in vivo. For instance, UDP-Glc concentration in the cytosol of spinach leaves was estimated as 1.7 mM [104], which should be compared with Arabidopsis UGPases’ Km values with UDP-Glc of 0.04–0.06 mM [51]. A similar estimation done for several metabolites in potato tubers [20] reveals that also other substrates of UGPase, in both directions of the reaction, are likely present at concentrations an order of magnitude higher than their corresponding Kms for the UGPase [51]. Similar conclusions can be drawn for most, if not all, of other NDP-sugar producing enzymes [50,64]. Thus, substrate availability is not a limiting factor in those reactions.

Even within the distinct metabolic domain of polysaccharide synthesis, there is a clear division of function for different nucleotides, with individual nucleotide pools regulating distinct metabolic processes (Figure 3). This is the case for (i) starch synthesis, which depends on adenylates [16], (ii) for Suc synthesis and the synthesis of cellulose and certain other components of cell walls, which are controlled by uridylates [50] and (iii) for the synthesis of polysaccharides from galactose, mannose or fucose, which is, at least partially, under the control of guanylates in plants [105]. There is even a role for cytidylates, which are needed for the activation of Kdo (Table 1 and Figure 3), a component of pectin rhamnogalacturonan II [38]. All these processes are controlled via AK/NDPK equilibria, and the displacement in these equilibria may direct metabolism towards the synthesis of a particular carbohydrate.

The equilibria established by NDP-sugar producing enzymes, under the overall control of AK/NDPK, are governed by various factors including the concentrations of substrates and products of the reactions, divalent ions, metabolic regulators and other factors. The differences in regulation of the enzymes linked to different nucleotides can provide a switch between corresponding metabolic pathways, e.g. AGPase is strongly inhibited by Pi which also suppresses the binding of PGA, the activator, while UGPase is not prone to PGA and Pi regulation [106]. In addition, the redox state can control AGPase [107] and UGPase [108] as well as some enzymes interconverting nucleotide sugars [109]. The existence of equilibria encompassing not only NDP-sugar forming enzymes, but also frequently enzymes preceding the pyrophosphorylase step, as e.g. for the conversion between Fru-6-P and UDP-Glc and/or ADP-Glc (Figure 1A), brings about even more players to the thermodynamic equilibrium concept.

Among factors affecting the local equilibria, a notable example is represented by PPi, the product of all pyrophosphorylase reactions. The PPi hydrolysis plays an important role in directing the reactions catalyzed by NDP-sugar pyrophosphorylases toward NDP-sugars, and prevents toxic effects of PPi, in particular, in the suppression of polysaccharide synthesis [110]. Both soluble pyrophosphatases (PPases) encoded by five genes in Arabidopsis and vacuolar proton pumping pyrophosphatase (H+-PPase) play a role in the maintenance of low concentrations of PPi [111]. Under hypoxic stress, the PPi can be produced in the pyruvate kinase/pyruvate, phosphate dikinase (PK/PPDK) substrate cycle, with its total balance of ADP + Pi → AMP + PPi. This maintains relatively stable levels of PPi in the conditions of ATP drop [112], and makes PPi an alternative energy currency [4]. The 100-fold increase in PPDK and several-fold increase in PK transcripts in rice under anoxia [113] supports the role of PPi in energy metabolism under stress, which also leads to feedback inhibition of polysaccharide synthesis due to the maintenance of PPi concentrations in the conditions of the drop of ATP and other NTPs. A role of PPi in the regulation of growth at different developmental stages via its effects on glycolysis and sugar synthesis has also been demonstrated [114].

Free magnesium (Mg2+) content, controlled in a given cell compartment by AK equilibrium [6,7,94], is essential for NDP-sugar production. For instance, UGPase from sorghum seedlings was found to use MgUTP and Glc-1-P in the forward direction of the reaction, and UDP-Glc and MgPPi as substrates in the reverse reaction, with Mg2+ stimulating the forward, but not reverse, reaction [115]. Magnesium complexed with PPi is also a natural substrate for pyrophosphatases, which use PPi produced by pyrophosphorylases [116]. Assuming that other pyrophosphorylases also utilize only Mg-bound NTP and Mg-bound PPi in their forward and reverse reactions, respectively, and that Mg2+ stimulates NDP-sugar formation, this would strongly suggest an indirect role of AK in polysaccharide formation. In this way, AK provides the constant load of the substrate, either directly (MgATP) or indirectly (MgUTP, MgGTP and MgCTP) by linking, via NDPK, with kinases of uridylate, guanylate and cytidylate metabolism, and controls Mg2+ content as an effector for pyrophosphorylases action.

Another example of Mg2+ involvement is its activation of SuSy toward UDP-Glc formation and inhibition of the reverse reaction [117]. A similar role in activating SuSy toward UDP-Glc (or ADP-Glc) formation was observed for phosphorylated (soluble) SuSy isozymes in comparison with non-phosphorylated (membrane-bound) SuSy isozymes [118], again suggesting an indirect role of AK/NDPK in controlling cytosolic sugar metabolism. The Mg2+ was also required for GDP-l-fucose pyrophosphorylase (FKGP, Table 1) activity in addition to its role in complexing GTP to form MgGTP, the true substrate of the reaction [119]. Increased Mg2+ also affected AGase (a phosphorylase) performance by changing its Keq toward ADP-Glc breakdown, and thus potentially affecting starch synthesis [70]. The change of Mg2+ concentration as a feedback of the equilibrium governed by AK, NDPK and other nucleoside kinases results in corresponding changes in internal Ca2+ since Ca2+ is chelated by nucleotides to a nearly the same extent as Mg2+. This triggers Ca-dependent signaling events in metabolism that in turn affect many processes including polysaccharide synthesis [120].

A simplified view of enzymatic reactions shown in Figure 3 is presented as a diagram in Figure 4. Generally, as a consequence of electron transport activity and the generation of membrane potential, ATP is synthesized via the oxidative and photosynthetic phosphorylation in mitochondria and chloroplasts, respectively. ATP is then used to generate other NTPs, which are equilibrated as NTPs, NDPs and NMPs by coupled reversible reactions of AK and NDPK. Various nucleoside phosphates are then used for de novo production of nucleotide sugars (see also Table 1). As AK is intimately involved in controlling Mg2+ content [3,6,7,92], the consequence of this is that it also indirectly regulates enzymes requiring Mg2+ for activity, e.g. those producing nucleotide sugars. From ATP to nucleotide sugar level all or most of the reactions can be considered as being in equilibrium, whereas the immediate reactions producing starch and cell wall polysaccharides are irreversible. This fulfills the major premise of the thermodynamic buffering concept, i.e. the direct link between reversible ‘buffering’ reactions (producing nucleotide sugars) and irreversible ‘engine’ reactions (producing polysaccharides from nucleotide sugars). Such a set-up is an important part of the metabolic control theory [12–14] and is computable [8], provided that the equilibrium constants of the reactions involved and the relevant metabolite contents in a given compartment are known. In the latter case, separate metabolomics data are required for chloroplasts and cytosol fractions, respectively (for starch formation), cytosol (cellulose and callose formation) and cytosol and Golgi apparatus, respectively (hemicellulose and pectin formation).

A simplified view of the links between nucleotide metabolome and polysaccharide formation in plants.

Figure 4.
A simplified view of the links between nucleotide metabolome and polysaccharide formation in plants.

Please note that the equilibration of nucleoside phosphates is carried out by AK in conjunction with NDPK as well as with guanylate kinase, UMP kinase and CMP kinase. Abbreviations: AK, adenylate kinase; ETC, electron transport chain (both in chloroplasts and mitochondria).

Figure 4.
A simplified view of the links between nucleotide metabolome and polysaccharide formation in plants.

Please note that the equilibration of nucleoside phosphates is carried out by AK in conjunction with NDPK as well as with guanylate kinase, UMP kinase and CMP kinase. Abbreviations: AK, adenylate kinase; ETC, electron transport chain (both in chloroplasts and mitochondria).

Conclusion

Thermodynamic buffering is a basic and common mechanism regulating the supply-demand balance in cell metabolism. It aims for the maintenance of the stability of non-equilibrium homeostatic fluxes in the system via the establishment of internal local equilibria. These local equilibria apply the ordering factor in the system and allow its stable operation resistant to external disturbances. The action of fast and reversible ‘buffering’ enzymes optimizes the function of slow and generally irreversible ‘engine’ enzymes which can be traced in different metabolic pathways and cycles.

In the current paper, we identified several pathways for polysaccharide formation in plants as confirming with basic premises of the thermodynamic buffering theory. The fast equilibrium reactions catalyzed mainly by pyrophosphorylases establish local equilibria of nucleotide sugars which include carbohydrate derivatives bound to adenylates, uridylates and guanylates. This results in the availability of ADP-, UDP- and GDP-bound sugars for their further conversion, via irreversible reactions of specific glycosyltransferases (‘engine’ enzymes), to di-, oligo- and polysaccharides and the possibility of a separate control of biosynthesis of carbohydrate polymers associated with different nucleotide pools. In addition, the equilibrium governed by AK, NDPK and other kinases not only provides nucleotide substrates for polysaccharide synthesis but establishes the concentration of Mg2+ (and other metal cations) that represents an important factor in the operation of various enzymes participating in sugar metabolism. Polysaccharide synthesis is an essential part of primary metabolism in plants that can now be computed/modeled within the thermodynamic buffering concept, provided that relevant compartment-targeted metabolome data are available.

Abbreviations

     
  • AGase

    ADP-Glc phosphorylase

  •  
  • AGPase

    ADP-Glc pyrophosphorylase

  •  
  • AK

    adenylate kinase

  •  
  • CalS

    callose synthase

  •  
  • CesA

    cellulose synthase

  •  
  • CKS

    CMP-Kdo synthetase

  •  
  • CMP-Kdo

    CMP-keto-deoxyoctulosonate

  •  
  • NDPK

    nucleoside-diphosphate kinase

  •  
  • NTPs

    nucleoside triphosphates

  •  
  • PGI

    phosphoglucoisomerase

  •  
  • PGM

    phosphoglucomutase

  •  
  • PK

    pyruvate kinase

  •  
  • PPDK

    pyruvate, phosphate dikinase

  •  
  • SPS

    Suc-P synthase

  •  
  • UAGPase

    UDP-GlcNAc (UDP-GalNAc) pyrophosphorylase

  •  
  • UDP-Glc

    UDP-d-glucose

  •  
  • UGPase

    UDP-Glc pyrophosphorylase

  •  
  • USPase

    UDP-sugar pyrophosphorylase

Author Contribution

Both authors have made substantial, direct and intellectual contribution to the work, and approved it for publication.

Funding

The research on the topics related to this manuscript was funded by the strategic fund from Umeå University (to L.A.K.) and by the Natural Sciences and Engineering Research Council of Canada (to A.U.I.).

Competing Interests

The authors declare that there are no competing interests associated with the manuscript.

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