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Molecular basis of the fructose-2,6-bisphosphatase reaction of PFKFB3: transition state and the C-terminal function.

Cavalier MC, Kim SG, Neau D, Lee YH
Proteins 80 1143-53 (2012)
Related entries: 3qpu, 3qpw

Cited: 16 times
EuropePMC logo PMID: 22275052

Abstract

The molecular basis of fructose-2,6-bisphosphatase (F-2,6-P(2)ase) of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFKFB) was investigated using the crystal structures of the human inducible form (PFKFB3) in a phospho-enzyme intermediate state (PFKFB3-P•F-6-P), in a transition state-analogous complex (PFKFB3•AlF(4)), and in a complex with pyrophosphate (PFKFB3•PP(i)) at resolutions of 2.45, 2.2, and 2.3 Å, respectively. Trapping the PFKFB3-P•F-6-P intermediate was achieved by flash cooling the crystal during the reaction, and the PFKFB3•AlF(4) and PFKFB3•PP(i) complexes were obtained by soaking. The PFKFB3•AlF(4) and PFKFB3•PP(i) complexes resulted in removing F-6-P from the catalytic pocket. With these structures, the structures of the Michaelis complex and the transition state were extrapolated. For both the PFKFB3-P formation and break down, the phosphoryl donor and the acceptor are located within ~5.1 Å, and the pivotal point 2-P is on the same line, suggesting an "in-line" transfer with a direct inversion of phosphate configuration. The geometry suggests that NE2 of His253 undergoes a nucleophilic attack to form a covalent N-P bond, breaking the 2O-P bond in the substrate. The resulting high reactivity of the leaving group, 2O of F-6-P, is neutralized by a proton donated by Glu322. Negative charges on the equatorial oxygen of the transient bipyramidal phosphorane formed during the transfer are stabilized by Arg252, His387, and Asn259. The C-terminal domain (residues 440-446) was rearranged in PFKFB3•PP(i), implying that this domain plays a critical role in binding of substrate to and release of product from the F-2,6-P(2) ase catalytic pocket. These findings provide a new insight into the understanding of the phosphoryl transfer reaction.

Articles - 3qpv mentioned but not cited (2)

  1. Molecular basis of the fructose-2,6-bisphosphatase reaction of PFKFB3: transition state and the C-terminal function. Cavalier MC, Kim SG, Neau D, Lee YH. Proteins 80 1143-1153 (2012)
  2. Intracellular BAPTA directly inhibits PFKFB3, thereby impeding mTORC1-driven Mcl-1 translation and killing MCL-1-addicted cancer cells. Sneyers F, Kerkhofs M, Speelman-Rooms F, Welkenhuyzen K, La Rovere R, Shemy A, Voet A, Eelen G, Dewerchin M, Tait SWG, Ghesquière B, Bootman MD, Bultynck G. Cell Death Dis 14 600 (2023)


Reviews citing this publication (3)

  1. Role of PFKFB3 and PFKFB4 in Cancer: Genetic Basis, Impact on Disease Development/Progression, and Potential as Therapeutic Targets. Kotowski K, Rosik J, Machaj F, Supplitt S, Wiczew D, Jabłońska K, Wiechec E, Ghavami S, Dzięgiel P. Cancers (Basel) 13 909 (2021)
  2. Fructose 2,6-Bisphosphate in Cancer Cell Metabolism. Bartrons R, Simon-Molas H, Rodríguez-García A, Castaño E, Navarro-Sabaté À, Manzano A, Martinez-Outschoorn UE. Front Oncol 8 331 (2018)
  3. The potential utility of PFKFB3 as a therapeutic target. Bartrons R, Rodríguez-García A, Simon-Molas H, Castaño E, Manzano A, Navarro-Sabaté À. Expert Opin. Ther. Targets 22 659-674 (2018)

Articles citing this publication (11)

  1. IKKβ promotes metabolic adaptation to glutamine deprivation via phosphorylation and inhibition of PFKFB3. Reid MA, Lowman XH, Pan M, Tran TQ, Warmoes MO, Ishak Gabra MB, Yang Y, Locasale JW, Kong M. Genes Dev. 30 1837-1851 (2016)
  2. Metabolic heterogeneity of idiopathic pulmonary fibrosis: a metabolomic study. Zhao YD, Yin L, Archer S, Lu C, Zhao G, Yao Y, Wu L, Hsin M, Waddell TK, Keshavjee S, Granton J, de Perrot M. BMJ Open Respir Res 4 e000183 (2017)
  3. Non-canonical roles of PFKFB3 in regulation of cell cycle through binding to CDK4. Jia W, Zhao X, Zhao L, Yan H, Li J, Yang H, Huang G, Liu J. Oncogene 37 1685-1698 (2018)
  4. KDM2A Targets PFKFB3 for Ubiquitylation to Inhibit the Proliferation and Angiogenesis of Multiple Myeloma Cells. Liu X, Li J, Wang Z, Meng J, Wang A, Zhao X, Xu Q, Cai Z, Hu Z. Front Oncol 11 653788 (2021)
  5. Pfkfb (6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase) isoforms display a tissue-specific and dynamic expression during Xenopus laevis development. Pegoraro C, Maczkowiak F, Monsoro-Burq AH. Gene Expr. Patterns 13 203-211 (2013)
  6. Metabolic regulation of calcium pumps in pancreatic cancer: role of phosphofructokinase-fructose-bisphosphatase-3 (PFKFB3). Richardson DA, Sritangos P, James AD, Sultan A, Bruce JIE. Cancer Metab 8 2 (2020)
  7. Glucose Drives Growth Factor-Independent Esophageal Cancer Proliferation via Phosphohistidine-Focal Adhesion Kinase Signaling. Zhang J, Gelman IH, Katsuta E, Liang Y, Wang X, Li J, Qu J, Yan L, Takabe K, Hochwald SN. Cell Mol Gastroenterol Hepatol 8 37-60 (2019)
  8. Hyperglycemia induces PFKFB3 overexpression and promotes malignant phenotype of breast cancer through RAS/MAPK activation. Cheng X, Jia X, Wang C, Zhou S, Chen J, Chen L, Chen J. World J Surg Oncol 21 112 (2023)
  9. Klotho increases antioxidant defenses in astrocytes and ubiquitin-proteasome activity in neurons. Orellana AM, Mazucanti CH, Dos Anjos LP, de Sá Lima L, Kawamoto EM, Scavone C. Sci Rep 13 15080 (2023)
  10. Mining anion-aromatic interactions in the Protein Data Bank. Kuzniak-Glanowska E, Glanowski M, Kurczab R, Bojarski AJ, Podgajny R. Chem Sci 13 3984-3998 (2022)
  11. Tuning PFKFB3 Bisphosphatase Activity Through Allosteric Interference. Macut H, Hu X, Tarantino D, Gilardoni E, Clerici F, Regazzoni L, Contini A, Pellegrino S, Luisa Gelmi M. Sci Rep 9 20333 (2019)


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