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Human UTP—glucose-1-phosphate uridylyltransferase isoform 1
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EC no.2.7.7.9
CAS no.9026-22-6
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UTP—glucose-1-phosphate uridylyltransferase also known as glucose-1-phosphate uridylyltransferase (or UDP–glucose pyrophosphorylase) is an enzyme associated with glycogenesis. It synthesizes UDP-glucose from glucose-1-phosphate and UTP; i.e.,

glucose-1-phosphate + UTP UDP-glucose + pyrophosphate

UTP—glucose-1-phosphate uridylyltransferase is an enzyme found in all three domains (bacteria, eukarya, and archaea) as it is a key player in carbohydrate metabolism. Its role in sugar metabolism has been studied extensively in plants in order to understand plant growth and increase agricultural production. Recently, human UDP-glucose pyrophosphorylase has been studied and crystallized, revealing a different type of regulation than other organisms previously studied. Its significance is derived from the many uses of UDP-glucose including galactose usage, glycogen synthesis, glycoprotein synthesis, and glycolipid synthesis.[1] [2][3]

Structure

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The structure of UTP—glucose-1-phosphate uridylyltransferase is significantly different between prokaryotes and eukaryotes, but within eukaryotes, the sequence and tertiary structure of the enzyme are quite conserved.[4] In many species, UTP—glucose-1-phosphate uridylyltransferase is found as a homopolymer consisting of identical subunits in a symmetrical quaternary structure.[5][6] The number of subunits varies across species: for instance, in E. coli, the enzyme is found as a tetramer, whereas in Burkholderia xenovorans, the enzyme is dimeric.[7][8] In humans, the enzyme is active as an octamer consisting of two tetramers stacked onto one another.[9]

In humans, each enzyme subunit contains several residues (L113, N251, and N328) that are highly conserved in eukaryotes. A Rossman fold motif participates in binding of the UTP nucleotide and a sugar-binding domain (residues T286–G293) coordinates with the glucose ring.[10] A missense mutation (G115D) in the region of the enzyme containing the active site (which is conserved in eukaryotes) causes a dramatic decrease in enzymatic activity in vitro.[11]

  • Crystal structure of UTP-glucose-1-phosphate uridylyltransferase from Burkholderia xenovorans
    Crystal structure of UTP-glucose-1-phosphate uridylyltransferase from Burkholderia xenovorans
  • Human UDP-glucose pyrophosphorylase isoform 1 subunit with UDP-glucose bound
    Human UDP-glucose pyrophosphorylase isoform 1 subunit with UDP-glucose bound

Examples

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Human genes encoding proteins with UTP—glucose-1-phosphate uridylyltransferase activity include two isoforms with molecular weights of 56.9 and 55.7 kDa, respectively.[12]

UDP–glucose pyrophosphorylase 1
Identifiers
SymbolUGP1
NCBI gene7359
HGNC12526
OMIM191750
Other data
EC number2.7.7.9
LocusChr. 1 q21-q22
UDP–glucose pyrophosphorylase 2
Identifiers
SymbolUGP2
NCBI gene7360
HGNC12527
OMIM191760
RefSeqNM_006759
UniProtQ16851
Other data
EC number2.7.7.9
LocusChr. 2 p14-p13
Search for
StructuresSwiss-model
DomainsInterPro

Function

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UTP—glucose-1-phosphate uridylyltransferase is ubiquitous in nature due to its important role in the generation of UDP-glucose, a central compound in carbohydrate metabolism. In plant leaves, UDP–glucose pyrophosphorylase is a key part of the sucrose biosynthesis pathway, supplying Uridine diphosphate glucose to Sucrose-phosphate synthase which converts UDP-glucose and D-fructose 6-phosphate into sucrose-6-phosphate.[13] It may also be partially responsible for the breakdown of sucrose in other tissues using UDP-glucose.

In higher animals, the enzyme is highly active in tissues involved in glycogenesis, including the liver and the muscles.[14] An exception is the brain, which has high levels of glycogen but low specific activity of UTP—glucose-1-phosphate uridylyltransferase.[15] In animal cells, UTP—glucose-1-phosphate uridylyltransferase is found predominantly in the cytoplasm.

UTP—glucose-1-phosphate uridylyltransferase is also required for galactose metabolism in animals and microorganisms. In galactose metabolism, the enzyme galactose 1-phosphate uridylyltransferase transfers a phosphate from UDP-glucose to galactose 1-phosphate to produce UDP-galactose, which is then converted to UDP-glucose.[16] Bacteria that have defective UTP—glucose-1-phosphate uridylyltransferase are unable to incorporate galactose into their cell walls.[17]

Mechanism

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In this enzyme’s primary reaction, the phosphate group on glucose-1-phosphate replaces the phosphoanhydride bond on UTP. This reaction is readily reversible and the Gibbs Free Energy is close to zero. However, under typical cellular conditions, inorganic pyrophosphatase quickly hydrolyzes the pyrophosphate product and drives the reaction forward by Le Chatelier's Principle.

UTP—glucose-1-phosphate uridylyltransferase uses an ordered sequential Bi Bi mechanism for both the forward and reverse reactions.[18]

Similar to other sugar nucleotidyltransferases, UTP—glucose-1-phosphate uridylyltransferase activity requires two divalent cations to stabilize the binding of negatively charged phosphate groups.[19] Magnesium typically serves in this role, but other ions such as manganese(II), cobalt(II), and nickel(II) can also substitute with a ~75% reduction in the optimal activity.[20] X-ray crystallography experiments have shown that one Mg2+ ion is coordinated by a phosphoryl oxygen on glucose 1-phosphate and by an α-phosphoryl oxygen on UTP.[21] In addition to stabilizing the negatively charged phosphates, Mg2+ is thought to orient the glucose 1-phosphate for nucleophilic attack of the α-phosphorus of UTP.[22]

Mechanism
UTP—glucose-1-phosphate uridylyltransferase reaction mechanism

Regulation

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Although functionally similar across species, UDP-glucose pyrophosphorylase has different structures and regulation mechanisms in different organisms.

Microorganisms

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In yeast, UTP—glucose-1-phosphate uridylyltransferase is regulated by phosphorylation by PAS kinase.[23] This phosphorylation is reversible and controls the partition of sugar flux towards glycogen and cell wall synthesis.

Plants

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UDP–glucose pyrophosphorylase in plants is regulated through oligomerization and possibly phosphorylation.[24] In barley, it has been shown that UDP-glucose pyrophosphorylase is only active in monomeric form but readily forms oligomers, suggesting that oligomerization may be a form of regulation of the enzyme. In rice, cold stress decreases N-glycosylation of the enzyme, which is thought to alter the enzyme's activity in response to cold.[25]

In Arabidopsis, there are two isozymes of UTP—glucose-1-phosphate uridylyltransferase: UGP1 and UGP2.[26] These two isozymes have almost identical activities and differ in only 32 amino acids, all of which are located on the outer surface of the protein away from the active site. These minor differences may allow for differential regulation of isozyme activity. UGP1 and UGP2 are differentially expressed in different parts of the plant. UGP1 expression is widely expressed in the majority of tissues while UGP2 is expressed primarily in flowers, suggesting that UGP1 is the major form of the enzyme and UGP2 serves an auxillary function. Indeed, UGP2 expression is increased in response to stressors such as phosphate deficiency, indicating that UGP2 probably functions as a backup to UGP1 when the plant is under environmental stress.

Animals

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The control of UTP—glucose-1-phosphate uridylyltransferase activity is primarily achieved by genetic means (i.e. regulation of enzyme synthesis). Similar to most enzymes, UTP—glucose-1-phosphate uridylyltransferase is inhibited by its product, UDP-glucose. However, the enzyme is not subject to significant allosteric control, which is logical given the widespread use of UDP-glucose in a variety of metabolic pathways.

Humans

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In humans, UDP-glucose pyrophosphorylase is active as an octamer.[9] This in turn lowers the activity of human UDP-glucose pyrophosphorylase compared to yeast UDP-glucose pyrophosphorylase. The difference in structure is potentially explained by the different UDP-glucose requirements of organisms with lower organisms such as yeast requiring more UDP-glucose than higher organisms such as mammals.[27] The enzyme's activity is also modified by O-glycosylation.[28] Similar to other mamallian species, there two different isoforms in humans that are produced by alternative splicing of the gene.[29][30][31] The isoforms differ by only 11 amino acids at the N-terminus and no significant differences in their functional activity have been identified.

Disease relevance

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In humans, galactosemia is a disorder that affects the development of newborns and children as they cannot metabolize the sugar galactose properly. It is speculated that overexpression of UDP-glucose pyrophosphorylase may relieve symptoms in humans with galactosemia.[32] The enzyme has also been found to be required for the biosynthesis of capsular polysaccharide, an important virulence factor of streptococcus pneumoniae, a bacterial cause of pneumonia, bronchitis, and other breathing issues.[33]

In cancer cells, which typically have high rates of glycolysis and decreased glycogen content, the activity of UTP—glucose-1-phosphate uridylyltransferase is often downregulated by up to 50-60% compared to normal cells[34]. The abnormally low activity of UTP—glucose-1-phosphate uridylyltransferase is due to decreased levels of the enzyme and the downregulation of other enzymes in the glycogenic pathway including glycogen synthase and phosphoglucomutase.

UTP—glucose-1-phosphate uridylyltransferase has been found to be an important virulence factor in a variety of pathogens including bacteria and protozoa.[35][36] As a result, the enzyme has attracted attention as a potential target for pharmaceuticals. However, in order to achieve specificity, the drugs must be designed to specifically target allosteric sites on the surface of the protein since the active site is highly conserved across species.[37]

References

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  1. ^ Sandhoff K, van Echten G, Schröder M, Schnabel D, Suzuki K (Aug 1992). "Metabolism of glycolipids: the role of glycolipid-binding proteins in the function and pathobiochemistry of lysosomes". Biochemical Society Transactions. 20 (3): 695–699. doi:10.1042/bst0200695. PMID 1426613.
  2. ^ Sandhoff K, van Echten G, Schröder M, Schnabel D, Suzuki K (Aug 1992). "Metabolism of glycolipids: the role of glycolipid-binding proteins in the function and pathobiochemistry of lysosomes". Biochemical Society Transactions. 20 (3): 695–699. doi:10.1042/bst0200695. PMID 1426613.
  3. ^ Alonso MD, Lomako J, Lomako WM, Whelan WJ (Sep 1995). "A new look at the biogenesis of glycogen". FASEB Journal. 9 (12): 1126–1137. PMID 7672505.
  4. ^ Führing, Jana Indra; Cramer, Johannes Thomas; Schneider, Julia; Baruch, Petra; Gerardy-Schahn, Rita; Fedorov, Roman (2015-04-10). "A Quaternary Mechanism Enables the Complex Biological Functions of Octameric Human UDP-glucose Pyrophosphorylase, a Key Enzyme in Cell Metabolism". Scientific Reports. 5 (1). doi:10.1038/srep09618. ISSN 2045-2322.
  5. ^ Kim, Hun; Choi, Jongkeun; Kim, Truc; Lokanath, Neratur K.; Ha, Sung Chul; Suh, Se Won; Hwang, Hye-Yeon; Kim, Kyeong Kyu (2010-04-01). "Structural basis for the reaction mechanism of UDP-glucose pyrophosphorylase". Molecules and Cells. 29 (4): 397–405. doi:10.1007/s10059-010-0047-6. ISSN 0219-1032. PMID 20238176.
  6. ^ Thoden, James B.; Holden, Hazel M. (2017-03-05). "Active site geometry of glucose-1-phosphate uridylyltransferase". Protein Science : A Publication of the Protein Society. 16 (7): 1379–1388. doi:10.1110/ps.072864707. ISSN 0961-8368. PMC 2206702. PMID 17567737.{{cite journal}}: CS1 maint: PMC format (link)
  7. ^ Thoden, James B.; Holden, Hazel M. (2017-03-06). "Active site geometry of glucose-1-phosphate uridylyltransferase". Protein Science : A Publication of the Protein Society. 16 (7): 1379–1388. doi:10.1110/ps.072864707. ISSN 0961-8368. PMC 2206702. PMID 17567737.{{cite journal}}: CS1 maint: PMC format (link)
  8. ^ Disease, Seattle Structural Genomics Center for Infectious. "Crystal structure of UDP-glucose pyrophosporylase / UTP-glucose-1-phosphate uridylyltransferase from Burkholderia xenovorans". to be published. doi:10.2210/pdb5j49/pdb.
  9. ^ a b Yu Q, Zheng X (Mar 2012). "The crystal structure of human UDP-glucose pyrophosphorylase reveals a latch effect that influences enzymatic activity". The Biochemical Journal. 442 (2): 283–291. doi:10.1042/BJ20111598. PMID 22132858.
  10. ^ Kleczkowski, Leszek A.; Geisler, Matt; Fitzek, Elisabeth; Wilczynska, Malgorzata (2011-11-01). "A common structural blueprint for plant UDP-sugar-producing pyrophosphorylases". The Biochemical Journal. 439 (3): 375–379. doi:10.1042/BJ20110730. ISSN 1470-8728. PMID 21992098.
  11. ^ Flores-Díaz, M.; Alape-Girón, A.; Persson, B.; Pollesello, P.; Moos, M.; von Eichel-Streiber, C.; Thelestam, M.; Florin, I. (1997-09-19). "Cellular UDP-glucose deficiency caused by a single point mutation in the UDP-glucose pyrophosphorylase gene". The Journal of Biological Chemistry. 272 (38): 23784–23791. ISSN 0021-9258. PMID 9295324.
  12. ^ "UGP2 - UTP--glucose-1-phosphate uridylyltransferase - Homo sapiens (Human) - UGP2 gene & protein". www.uniprot.org. Retrieved 2017-03-06.
  13. ^ Mendicino J (Dec 1960). "Sucrose phosphate synthesis in wheat germ and green leaves". The Journal of Biological Chemistry. 235: 3347–3352. PMID 13769376.
  14. ^ Turnquist, Richard L.; Hansen, R. Gaurth (1973-01-01). Boyer, Paul D. (ed.). The Enzymes. Group Transfer Part A: Nucleotidyl Transfer Nucleosidyl Transfer Acyl Transfer Phosphoryl Transfer. Vol. 8. Academic Press. pp. 51–71.
  15. ^ Villar-Palasi, C.; Larner, J. (1960-03-25). "Insulin-mediated effect on the activity of UDPG-glycogen transglucosylase of muscle". Biochimica Et Biophysica Acta. 39: 171–173. ISSN 0006-3002. PMID 13842294.
  16. ^ Bosch, Annet M. (2006-08-01). "Classical galactosaemia revisited". Journal of Inherited Metabolic Disease. 29 (4): 516–525. doi:10.1007/s10545-006-0382-0. ISSN 1573-2665. PMID 16838075.
  17. ^ Sundararajan, T. A.; Rapin, A. M.; Kalckar, H. M. (1962-12-15). "Biochemical observations on E. coli mutants defective in uridine diphosphoglucose". Proceedings of the National Academy of Sciences of the United States of America. 48: 2187–2193. ISSN 0027-8424. PMC 221142. PMID 13979281.
  18. ^ Tsuboi, K. K.; Fukunaga, K.; Petricciani, J. C. (1969-02-10). "Purification and specific kinetic properties of erythrocyte uridine diphosphate glucose pyrophosphorylase". The Journal of Biological Chemistry. 244 (3): 1008–1015. ISSN 0021-9258. PMID 5782905.
  19. ^ Zea, Corbin J; Camci-Unal, Gulden; Pohl, Nicola L (2008-07-15). "Thermodynamics of binding of divalent magnesium and manganese to uridine phosphates: implications for diabetes-related hypomagnesaemia and carbohydrate biocatalysis". Chemistry Central Journal. 2 (1). doi:10.1186/1752-153x-2-15. ISSN 1752-153X.{{cite journal}}: CS1 maint: unflagged free DOI (link)
  20. ^ Gustafson, G. L.; Gander, J. E. (1972-03-10). "Uridine Diphosphate Glucose Pyrophosphorylase from Sorghum vulgare PURIFICATION AND KINETIC PROPERTIES". Journal of Biological Chemistry. 247 (5): 1387–1397. ISSN 0021-9258. PMID 5012314.
  21. ^ Thoden, James B.; Holden, Hazel M. (2017-03-06). "Active site geometry of glucose-1-phosphate uridylyltransferase". Protein Science : A Publication of the Protein Society. 16 (7): 1379–1388. doi:10.1110/ps.072864707. ISSN 0961-8368. PMC 2206702. PMID 17567737.{{cite journal}}: CS1 maint: PMC format (link)
  22. ^ Sivaraman, J.; Sauvé, Véronique; Matte, Allan; Cygler, Miroslaw (2002-11-15). "Crystal structure of Escherichia coli glucose-1-phosphate thymidylyltransferase (RffH) complexed with dTTP and Mg2+". The Journal of Biological Chemistry. 277 (46): 44214–44219. doi:10.1074/jbc.M206932200. ISSN 0021-9258. PMID 12171937.{{cite journal}}: CS1 maint: unflagged free DOI (link)
  23. ^ Rutter, Jared; Probst, Brandon L.; McKnight, Steven L. (2002-10-04). "Coordinate regulation of sugar flux and translation by PAS kinase". Cell. 111 (1): 17–28. ISSN 0092-8674. PMID 12372297.
  24. ^ Kleczkowski LA, Geisler M, Ciereszko I, Johansson H (Mar 2004). "UDP-glucose pyrophosphorylase. An old protein with new tricks". Plant Physiology. 134 (3): 912–918. doi:10.1104/pp.103.036053. PMID 15020755.
  25. ^ Komatsu, S.; Yamada, E.; Furukawa, K. (2008-02-16). "Cold stress changes the concanavalin A-positive glycosylation pattern of proteins expressed in the basal parts of rice leaf sheaths". Amino Acids. 36 (1): 115–123. doi:10.1007/s00726-008-0039-4. ISSN 0939-4451.
  26. ^ Meng, M.; Geisler, M.; Johansson, H.; Harholt, J.; Scheller, H. V.; Mellerowicz, E. J.; Kleczkowski, L. A. (2009-05-01). "UDP-Glucose Pyrophosphorylase is not Rate Limiting, but is Essential in Arabidopsis". Plant and Cell Physiology. 50 (5): 998–1011. doi:10.1093/pcp/pcp052. ISSN 0032-0781.
  27. ^ Yu Q, Zheng X (Mar 2012). "The crystal structure of human UDP-glucose pyrophosphorylase reveals a latch effect that influences enzymatic activity". The Biochemical Journal. 442 (2): 283–291. doi:10.1042/BJ20111598. PMID 22132858.
  28. ^ Wells, Lance; Hart, Gerald W. (2003-07-03). "O-GlcNAc turns twenty: functional implications for post-translational modification of nuclear and cytosolic proteins with a sugar". FEBS Letters. 546 (1): 154–158. doi:10.1016/S0014-5793(03)00641-0. ISSN 1873-3468.
  29. ^ Führing, Jana; Damerow, Sebastian; Fedorov, Roman; Schneider, Julia; Münster-Kühnel, Anja-Katharina; Gerardy-Schahn, Rita (2013-04-01). "Octamerization is essential for enzymatic function of human UDP-glucose pyrophosphorylase". Glycobiology. 23 (4): 426–437. doi:10.1093/glycob/cws217. ISSN 1460-2423. PMID 23254995.
  30. ^ Duggleby, R. G.; Chao, Y. C.; Huang, J. G.; Peng, H. L.; Chang, H. Y. (1996-01-15). "Sequence differences between human muscle and liver cDNAs for UDPglucose pyrophosphorylase and kinetic properties of the recombinant enzymes expressed in Escherichia coli". European Journal of Biochemistry. 235 (1–2): 173–179. ISSN 0014-2956. PMID 8631325.
  31. ^ "UGP2 - UTP--glucose-1-phosphate uridylyltransferase - Homo sapiens (Human) - UGP2 gene & protein". www.uniprot.org. Retrieved 2017-03-05.
  32. ^ Lai K, Elsas LJ (May 2000). "Overexpression of human UDP-glucose pyrophosphorylase rescues galactose-1-phosphate uridyltransferase-deficient yeast". Biochemical and Biophysical Research Communications. 271 (2): 392–400. doi:10.1006/bbrc.2000.2629. PMID 10799308.
  33. ^ Bonofiglio L, García E, Mollerach M (Oct 2005). "Biochemical characterization of the pneumococcal glucose 1-phosphate uridylyltransferase (GalU) essential for capsule biosynthesis". Current Microbiology. 51 (4): 217–221. doi:10.1007/s00284-005-4466-0. PMID 16132460.
  34. ^ Nigam, Vijai N.; MacDonald, Helen L.; Cantero, A. (1962-02-01). "Limiting Factors for Glycogen Storage in Tumors". Cancer Research. 22 (2): 131–138. ISSN 0008-5472. PMID 14479721.
  35. ^ Jiang, Sin-Sien; Lin, Tzu-Yi; Wang, Won-Bo; Liu, Ming-Che; Hsueh, Po-Ren; Liaw, Shwu-Jen (2010-05-01). "Characterization of UDP-glucose dehydrogenase and UDP-glucose pyrophosphorylase mutants of Proteus mirabilis: defectiveness in polymyxin B resistance, swarming, and virulence". Antimicrobial Agents and Chemotherapy. 54 (5): 2000–2009. doi:10.1128/AAC.01384-09. ISSN 1098-6596. PMC 2863647. PMID 20160049.{{cite journal}}: CS1 maint: PMC format (link)
  36. ^ Klein, Kathryn A.; Fukuto, Hana S.; Pelletier, Mark; Romanov, Galina; Grabenstein, Jens P.; Palmer, Lance E.; Ernst, Robert; Bliska, James B. (2012-02-01). "A transposon site hybridization screen identifies galU and wecBC as important for survival of Yersinia pestis in murine macrophages". Journal of Bacteriology. 194 (3): 653–662. doi:10.1128/JB.06237-11. ISSN 1098-5530. PMC 3264090. PMID 22139502.{{cite journal}}: CS1 maint: PMC format (link)
  37. ^ Führing, Jana; Damerow, Sebastian; Fedorov, Roman; Schneider, Julia; Münster-Kühnel, Anja-Katharina; Gerardy-Schahn, Rita (2013-04-01). "Octamerization is essential for enzymatic function of human UDP-glucose pyrophosphorylase". Glycobiology. 23 (4): 426–437. doi:10.1093/glycob/cws217. ISSN 1460-2423. PMID 23254995.
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