Plant Nucleotide Metabolism. Hiroshi Ashihara
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ADP + 3-Phospho-D-glyceroyl phosphate → ATP + 3-Phospho-D-glycerate
3.3.2 Specific Nucleoside-Monophosphate Kinases
In addition to the non-specific nucleoside-monophosphate kinase mentioned above, there are at least three nucleoside-monophosphate kinases which contribute to the conversion of NMP to NDP. They are adenylate (AMP) kinase (EC 2.7.4.3), guanylate (GMP) kinase (EC 2.7.4.8) and cytidylate (UMP/cytidine-5′-monophosphate [CMP]) kinase (EC 2.7.4.14). It has been reported that genes encoding adenylate kinase show high sequence homology with those encoding UMP-CMP kinase (Fukami-Kobayashi et al. 1996). The equilibrium constants of these enzymes are ∼1, so the reaction is reversible. Adenylate kinase is ubiquitous and is found in different subcellular locations, including the cytosol, mitochondria, and plastids of plants. The reactions of each enzyme, and associated references, are presented in Table 3.1.
3.4 Conversion of Nucleoside Diphosphates to Nucleoside Triphosphates
Phosphorylation of nucleoside diphosphates, especially ADP to ATP, is performed by two different types of reaction. One, ADP + Pi → ATP, is the electron transfer system of respiration and photosynthesis. The other is reactions involving substrate level phosphorylation, which is the formation of ATP or GTP by the direct transfer of a phosphoryl group to ADP or GDP from another phosphorylated metabolite, such as phosphoenolpyruvate.
3.4.1 ATP Synthesis by Electron Transfer Systems
The conversion of ADP to ATP is performed by phosphorylation linked to the electron transport chains of respiration and photosynthesis (Karp 2013; Niyogi et al. 2015). Briefly, in oxidative phosphorylation, electrons are derived from substrates that enter the TCA (tricarboxylic acid) cycle (aka the citrate or Krebs cycle) (Figure 3.2), whereas in photophosphorylation, they are furnished by chlorophyll in the presence of light. ATP synthesis is coupled to the subsequent successive oxidation–reduction of members of electron transport chains, which pass electron pairs through stages of successively lower potential energy until they reach oxygen, the terminal electron sink. ATP synthase (H+-transporting two-sector ATPase, EC 3.6.3.14) is an enzyme that creates ATP. It is formed from ADP and Pi (Table 3.1).
Figure 3.2 The tricarboxylic acid (Krebs) cycle in plants. (1) Pyruvate dehydrogenase; (2) citrate synthase; (3) aconitase; (4) isocitrate dehydrogenase; (5) 2-oxoglutarate dehydrogenase; (6) succinyl-CoA synthetase; (7) succinate dehydrogenase; (8) fumarase; (9) malate dehydrogenase.
3.4.2 Substrate-Level ATP Synthesis
Substrate-level phosphorylation of ADP is performed by the direct transfer of a phosphoryl group to ADP from another phosphorylated compound (Table 3.1). In the TCA cycle in plants, substrate-level phosphorylation occurs mainly in the cytosol or chloroplasts during glycolysis and in mitochondria.
Substrate-level phosphorylation in glycolysis occurs via two steps, 7 and 10, in Figure 3.3. After the conversion of glyceraldehyde-3-phosphate to glycerate-1,3-bisphosphate, catalysed by glyceraldehyde-3-phosphate dehydrogenase using Pi and NAD+ (step 6), ATP is formed by the dephosphorylation of glycerate-1,3-bisphosphate (step 7). The second substrate-level phosphorylation is catalysed by pyruvate kinase (step 10) and ATP is produced from the phosphate group of phosphoenolpyruvate. The hexose molecule is split into two three-carbon molecules, via an aldolase reaction (step 4). Hexose is converted to fructose-1,6-bisphosphate by hexokinase (step 1) and phosphofructokinase (step 3a), in the process consuming 2 mol and producing 4 mol of ATP in steps 7 and 10. Thus, there is net production of 2 mol of ATP. The ATP-consuming and ATP-producing reactions of glycolysis are illustrated in Figure 3.4. If the conversion is performed by hexokinase (step 1) and pyrophosphate (PPi)-dependent phosphofructokinase (step 3b), net production of ATP is 3 mol from 1 mol of hexose. Two mol of NADH produced by glycolysis can be used in oxidative phosphorylation in mitochondria to generate more ATP (Zeeman 2015).
A further substrate-level phosphorylation occurs in the TCA cycle. In the matrix of the mitochondria, a substrate-level phosphorylation occurs at the succinate-CoA ligase reaction step (step 6 in Fig. 3.2). In contrast to animals, in plants ATP-specific succinyl-CoA synthetase (EC 6.2.1.5), but not GTP-succinyl-CoA synthetase (EC 6.2.1.4), acts as an enzyme in the TCA cycle (see Figure 3.4). This enzyme produces ATP from ADP accompanied by degradation of succinyl-CoA (Table 3.1 and Figure 3.4c). For further details of this topic, readers can refer to comprehensive plant biochemistry text books such as Bowsher et al. (2012) and Buchanan et al. (2015).
3.4.3 Nucleoside-Diphosphate Kinase
Using cellular ATP, purine and pyrimidine nucleoside triphosphate are produced by nucleoside-diphosphate kinase (EC 2.7.4.6). The enzyme catalyses the reaction: nucleoside diphosphate + ATP ↔ nucleoside triphosphate + ADP. Substrate specificity is broad. All purine and pyrimidine nucleoside diphosphates can act as acceptors, while not only ATP, but also any ribonucleoside- and deoxyribonucleoside triphosphate, can act as a donor. This enzyme contributes principally to the formation of nucleoside triphosphates, such as GTP, uridine-triphosphate (UTP) and cytidine-triphosphate (CTP), from the respective nucleoside-diphosphates. Nucleoside-diphosphate kinase occurs ubiquitously in animals, plants, and bacteria. Recent molecular genetic studies suggest that nucleoside-diphosphate kinases are major players in the synthesis of macromolecules since they provide the triphosphates used for cell anabolism, including nucleic acid synthesis, CTP for lipid synthesis, UTP for polysaccharide synthesis, and GTP for protein elongation, signal transduction, and microtubule polymerization. Five nucleoside-diphosphate kinase genes have been detected in Arabidopsis thaliana and rice (Oryza sativa). Cytosolic (type I) and chloroplast (type II) enzymes are involved in metabolism, growth, and stress responses and in photosynthetic development and oxidative stress management, respectively. Type III enzymes are located in mitochondria and chloroplasts and are involved principally in energy metabolism. The subcellular localization and precise function of the novel type IV enzyme has not as yet been determined (Dorion and Rivoal 2015).
Figure 3.3 Glycolysis