Báo cáo khoa học: Pentose phosphates in nucleoside interconversion and catabolism

Pentose Phosphates in Nucleoside Interconversion and Catabolism

Tác giả: Maria G. Tozzi, Marcella Camici, Laura Mascia, Francesco Sgarrella, Piero L. Ipata

Lĩnh vực: Sinh hóa

Nội dung tài liệu: Bài tổng quan này tập trung vào sự chuyển đổi và dị hóa của các pentose phosphate, các hợp chất quan trọng trong chuyển hóa nucleoside. Tài liệu làm sáng tỏ các cơ chế phân tử liên quan đến việc chuyển hóa ribose-1-phosphate và deoxyribose-1-phosphate, vai trò của chúng trong quá trình tổng hợp nucleotide mới và quá trình “giải cứu” (salvage). Bài viết cũng thảo luận về cách các pentose phosphate, bắt nguồn từ quá trình phân hủy nucleoside, có thể được tái chế hoặc dị hóa thành nguồn carbon và năng lượng. Đặc biệt, nghiên cứu xem xét sự tương tác giữa các con đường trao đổi chất purine và pyrimidine, cũng như vai trò của nucleoside phosphorylase trong việc vận chuyển đường và các hợp chất dị hóa.

Mục lục chi tiết:

• Review Article: Pentose phosphates in nucleoside interconversion and catabolism
• Keywords
• Correspondence
• Pentose phosphates are either synthesized through the oxidative branch of the pentose phosphate pathway, or are supplied by nucleoside phosphorylases.
• The two main pentose phosphates, ribose-5-phosphate and ribose-1-phosphate, are readily interconverted by the action of phosphopentomutase.
• Ribose-5-phosphate is the direct precursor of 5-phosphoribosyl-1-pyrophosphate, for both de novo and ‘salvage’ synthesis of nucleotides.
• Phosphorolysis of deoxyribonucleosides is the main source of deoxyribose phosphates, which are interconvertible, through the action of phosphopentomutase.
• The pentose moiety of all nucleosides can serve as a carbon and energy source.
• During the past decade, extensive advances have been made in elucidating the pathways by which the pentose phosphates, arising from nucleoside phosphorolysis, are either recycled, without opening of their furanosidic ring, or catabolized as a carbon and energy source.
• We review herein the experimental knowledge on the molecular mechanisms by which (a) ribose-1-phosphate, produced by purine nucleoside phosphorylase acting catabolically, is either anabolized for pyrimidine salvage and 5-fluorouracil activation, with uridine phosphorylase acting anabolically, or recycled for nucleoside and base interconversion; (b) the nucleosides can be regarded, both in bacteria and in eukaryotic cells, as carriers of sugars, that are made available though the action of nucleoside phosphorylases.
• In bacteria, catabolism of nucleosides, when suitable carbon and energy sources are not available, is accomplished by a battery of nucleoside transporters and of inducible catabolic enzymes for purine and pyrimidine nucleosides and for pentose phosphates.
• In eukaryotic cells, the modulation of pentose phosphate production by nucleoside catabolism seems to be affected by developmental and physiological factors on enzyme levels.
• Pentose phosphates are heterocyclic, five-membered, oxygen-containing phosphorylated ring structures, with ribose-5-phosphate (Rib-5-P) and 2-deoxyribose-5-phosphate (deoxyRib-5-P) being basal structures of ribonucleotides and deoxyribonucleotides, respectively, and 5-phosphoribosyl-1-pyrophosphate (PRPP) the common precursor of both de novo and ‘salvage’ synthesis of nucleotides.
• Two main pathways are involved in pentose phosphate biosynthesis (Fig. 1).
• In the oxidative branch of the pentose phosphate pathway, Rib-5-P is generated from glucose-6-phosphate.
• In the phosphorylase-mediated pathway, deoxyribose-1-phosphate (deoxyRib-1-P) and ribose-1-phosphate (Rib-1-P) are supplied by various nucleoside phosphorylases, such as thymidine phosphorylase, uridine phosphorylase (UPase) and purine nucleoside phosphorylase (PNP).
• PNP deficiency causes a clinical syndrome of severe combined immunodeficiency, indistinguishable from that of adenosine deaminase deficiency [2,3].
• Rib-5-P may also be formed from free ribose by the action of ribokinase.
• The enzyme from Escherichia coli has been crystallized and its genetic regulation extensively studied in bacteria [4-8].
• However, the phosphorylation of free ribose by ribokinase is a less investigated pathway in mammals, even though its involvement in the elevation of PRPP, following ribose administration as a metabolic supplement for the heart and central nervous system, has been demonstrated [9,10].
• The reader is referred to the numerous excellent reviews covering the different aspects of nucleoside and nucleobase metabolism [11-13].
• This article focuses on the direct link between the ribose moiety of nucleosides and central carbon metabolism.
• Pentose phosphates in nucleoside interconversion
• PNP and UPase-mediated ribose transfer
• The equilibrium of PNP-catalysed reactions is thermodynamically in favour of nucleoside synthesis [1,14].
• Nevertheless, it is generally accepted that in vivo inosine and guanosine phosphorolysis is favoured (a) because the intracellular concentration of P₁ is higher than that of nucleosides [11] and (b) as a result of the coupling of liberated hypoxanthine and guanine with hypoxanthine-guanine phosphoribosyl transferase (HPRT) and, in certain tissues, xanthine oxidase or guanase, respectively, the equilibrium of the PNP reaction is shifted towards Rib-1-P accumulation (Fig. 2).
• Another important factor is the absence in mammals of any kinase acting on inosine and guanosine [15-17], which further favours the channelling of purine nucleosides towards phosphorolysis.
• Interestingly, purine ribonucleoside kinases are also absent in Lactococcus lactis, hence the only pathway for purine nucleoside salvage in this bacterium is through phosphorolytic cleavage by PNP to the free nucleobase and Rib-1-P [13].
• We can reasonably assume that in vivo PNP acts catabolically, leading to pentose phosphate formation for its further utilization in cell metabolism.
• A different metabolic situation may be envisaged for UPase.
• The homeostasis of uridine, which regulates several physiological and pathological processes [18], is maintained by the relative activities of two enzymes: the UTP-CTP inhibited uridine kinase [19,20] and UPase.
• It has long been assumed that UPase, in analogy to PNP, acts catabolically, even though in 1985 Schwartz et al. [21] gave convincing evidence for its anabolic role in 5-fluouracil (5-FU) activation to cytotoxic compounds.
• More recent in vitro experiments have established that indeed UPase may catalyse the Rib-1-P-mediated ribosylation of 5-FU and uracil, even in the presence of excess P₁ [20,22].
• In normal rat tissues and in PC12 cells, this process, called the ‘Rib-1-P pathway’, predominates over the one-step ‘PRPP pathway’, as catalysed by orotate phosphoribosyltransferase, and represents the only known way for salvaging uracil [20,23].
• Cao et al. [24] have developed a UPase gene knockout embryonic stem cell model and have shown that the disruption of UPase activity leads to a 10-fold increase in the 5-FU 50% inhibitory concentration (IC50), and to a two to threefold reduction in its incorporation into nucleic acids.
• At least in rat brain this ‘UPase-mediated anabolism’ (Fig. 2) is favoured because (a) degradation of uracil to ẞ-alanine, which would drive uridine phosphorolysis, is absent in the central nervous system (CNS) [14,25], (b) multiple consecutive phosphorylations of uridine by the ubiquitous uridine kinase and nucleoside mono- and diphosphokinases drive the Rib-1-P-mediated uracil and 5-FU ribosylation, and (c) the absence of uracil phosphoribosyltransferase in mammals [26] further channels Rib-1-P towards 5-FU and uracil ribosylation.
• We can therefore assume that the Rib-1-P produced by inosine phosphorolysis may, in part, become a substrate for 5-FU activation and for uracil salvage, thus establishing a metabolic link between purine and pyrimidine salvage synthesis (Fig. 2).
• In bacterial systems, whether UPase can be used anabolically for uptake of uracil without any ribose donors added may be determined in mutants lacking uracil phosphoribosyltransferase (upp pyr mutants) [13].
• In L. lactis, the low concentration of Rib-1-P makes the ribonucleoside synthesis unfavourable.
• Thus, in an upp pyr mutant, the irreversibility of UPase was shown by the inability of uracil to satisfy the pyrimidine requirement [27].
• However, when supplied with a purine nucleoside as a source of Rib-1-P, the uracil analogue, 5-FU, is converted to 5-fluorouridine [28].
• The inability to utilize uracil through UPase is also found in enteric bacteria [29].
• Usually wild-type bacteria, including Gram-positive bacteria, are unable to anabolize thymine.
• However, thymine-requiring mutants of E. coli and Salmonella typhimurium can deoxyribosylate thymine to thymidine by thymidine phosphorylase, because their deoxyRib-1-P pools are high [30].
• In these mutants, deoxyUTP accumulates and is broken down to deoxyuridine, which again is cleaved by thymidine phosphorylase to uracil and deoxyRib-1-P.
• The PNP-mediated ribose transfer from a nucleoside to a base analogue, with potential antiviral or antineoplastic activity, has been widely used for the in vitro synthesis of novel nucleoside analogues.
• Alternatively, a nucleoside modified in its ribose moiety may be used to obtain a new nucleoside analogue, modified in its pentose ring.
• The utility of this procedure was documented by Krenitski et al. in 1981 [31].
• Since then, a large variety of new nucleoside analogues have been enzymatically synthesized.
• We refer to the excellent review of Bzowska et al. [1] for furthering the principles and techniques related to this important field of applied enzymology.
• The recent introduction of thermostable phosphorylases isolated from Sulfolobus solfataricus and Pyrococcus furiosus [32,33] might offer a promising improvement.
• Rib-1-P recycling
• During the course of experiments designed to isolate deoxyRib-1-P formed by the reversible enzymatic phosphorolysis of deoxyguanosine catalysed by PNP, in 1952 Friedkin tried to increase the yield of deoxy-Rib-1-P by coupling deoxyguanosine phosphorolysis with the irreversible guanine deamination, catalysed by guanase [34].
• In theory, for each mole of deoxyguanosine undergoing phosphorolysis, one mole of xanthine and one mole of deoxyRib-1-P should also be formed.
• However, both xanthine and deoxyRib-1-P unexpectedly disappeared.
• This observation led to the isolation of deoxyxanthosine, a hitherto-undescribed deoxynucleoside, which was formed by deoxyribosylation of xanthine, catalysed by PNP.
• The sum of the three above-reported reactions is the hydrolytical deamination of deoxyguanosine, in the absence of a specific deoxyguanosine deaminase.
• Years later, an enzyme system, catalysing the apparent deamination of guanosine to xanthosine, was reconstituted in vitro, using commercial PNP and guanase [14].
• In this system, xanthine, after reaching a maximal value, decreased consistently in parallel with the increase of xanthosine.
• Moreover, replacement of Pi with arsenate, hindering the formation of Rib-1-P, prevented the formation of xanthosine, but not that of guanine and xanthine.
• The Rib-1-P recycling for guanosine deamination is operative in rat liver [14,34] and brain [35], and might be responsible for the presence of xanthosine in human serum and tissues [36].
• In both the ‘UPase-mediated Rib-1-P anabolism’ and the ‘Rib-1-P recycling for nucleoside and base interconversion’, the ribose moiety of Rib-1-P, produced by the action of PNP, is transferred to a nucleobase.
• Nevertheless, the two processes are metabolically different.
• In the first, the net reaction is the transfer of ribose from a nucleoside to a base, with Rib-1-P acting as a form of activated ribose.
• In the second, the net reaction is the hydrolytic deamination of guanosine, with Rib-1-P acting catalytically [14] (Fig. 2).
• A similar Rib-1-P recycling system is operative in Bacillus cereus [37].
• This organism does not possess any adenine deaminase, yet it can quantitatively mobilize the amino group of adenine for biosynthetic reactions by catalysing the ribosylation of adenine by adenosine phosphorylase, an enzyme distinct from PNP [38], followed by adenosine deamination and inosine phosphorolysis.
• Alternatively, adenosine can be phosphorylated to AMP by adenosine kinase [39].
• Rib-1-P recycling also occurs in E. coli and L. lactis.
• In these organisms, free adenine can serve as the sole purine source.
• Adenine is converted into adenosine, and then into inosine and hypoxanthine using the Rib-1-P recycling process, and after conversion of hypoxanthine to inosine-5′-mono-phosphate (IMP), these reactions in summary result in the conversion of adenine into IMP, which serves as a precursor for guanosine-5′-monophosphate (GMP) synthesis [13].
• Mammals do not possess any adenosine phosphorylase activity, therefore they cannot carry out these kinds of Rib-1-P recycling.
• N-deoxyribosyltransferases
• Contrary to the ribose moiety of inosine, which must be transformed by PNP into free Rib-1-P in order to be transferred to a nucleobase, the deoxyribose moiety of deoxyinosine can be transferred to a nucleobase acceptor by a single enzyme protein, the N-deoxyribosyltransferase, without the intermediate formation of free deoxyRib-1-P.
• The glycosyl transfer is stereospecific, in that only the ẞ-anomer of the deoxynucleoside is formed.
• The enzyme, first discovered by McNutt in 1952 [40], is present in Lactobacillus species, which are devoid of nucleoside phosphorylases and hence cannot degrade or synthesize deoxyribonucleosides phosphorolytically.
• As they also often have a growth requirement for deoxynucleosides, it is important that these compounds are not degraded when present in the medium.
• The presence of the N-deoxyribosyltransferase and all four nucleobases found in DNA and just one deoxynucleoside ensures a supply of all four deoxynucleotides, because these bacteria possess deoxynucleoside kinase activities.
• The genes encoding two distinct N-deoxyribosyltransferases have been isolated by Kaminski [41].
• The wide specificity of the two transferases for deoxynucleoside donors and base acceptors made it possible to synthesize a large number of deoxynucleoside analogues with potential antiviral and antineoplastic activity [42].
• Pentose phosphates as a carbon and an energy source
• As this section is devoted to the catabolism of the ribose moiety of both intracellular and extracellular nucleotides, an introduction on the reactions involved in this pathway and on the enzymes catalysing these reactions appears to be necessary (Fig. 3).
• Nucleoside phosphorylases play a key role in the utilization of nucleosides [1].
• Based on their structural properties, nucleoside phosphorylases have been classified into two families: NP-I and NP-II.
• The NP-I family includes homotrimeric and homohexameric enzymes from both prokaryotes and eukaryotes acting on inosine, guanosine, adenosine and uridine.
• The NP-II family includes homodimeric proteins structurally unrelated to the NP-I family, such as bacterial pyrimidine phosphorylases and eukaryotic thymidine phosphorylase [43].
• This enzyme was shown to be identical to the platelet-derived endothelial cell growth factor, a protein known to possess chemotactic activity in vitro and angiogenic activity in vivo [44].
• However, stimulation of endothelial cell proliferation was soon after ascribed to the deoxyribose arising from the intracellular breakdown of thymidine, rather than to an intrinsic property of thymidine phosphorylase [45].
• Phosphopentomutase catalyses the reversible reaction between Rib-1-P and Rib-5-P and between deoxyRib-1-P and deoxyRib-5-P.
• The enzyme has been extensively studied in bacteria [46-48].
• Among eukaryotes, phosphopentomutase activity has been detected in rabbit tissues [49], human leukemic cells [50], human erythrocytes [51] and in a cell line derived from the human amnion epithelium (WISH) [52], and has been purified from rat liver [53].
• The key enzyme for the catabolism of the pentose moiety of deoxyribonucleosides is deoxyriboaldolase, which cleaves deoxyRib-5-P into acetaldehyde and glyceraldehyde 3-P.
• Bacterial deoxy