title: Structures of human phosphofructokinase-1 and atomic basis of cancer-associated mutations.
abstract: Phosphofructokinase-1 (PFK1), the 'gatekeeper' of glycolysis, catalyses the committed step of the glycolytic pathway by converting fructose-6-phosphate to fructose-1,6-bisphosphate. Allosteric activation and inhibition of PFK1 by over ten metabolites and in response to hormonal signalling fine-tune glycolytic flux to meet energy requirements. Mutations inhibiting PFK1 activity cause glycogen storage disease type VII, also known as Tarui disease, and mice deficient in muscle PFK1 have decreased fat stores. Additionally, PFK1 is proposed to have important roles in metabolic reprogramming in cancer. Despite its critical role in glucose flux, the biologically relevant crystal structure of the mammalian PFK1 tetramer has not been determined. Here we report the first structures of the mammalian PFK1 tetramer, for the human platelet isoform (PFKP), in complex with ATP-Mg(2+) and ADP at 3.1 and 3.4 Å, respectively. The structures reveal substantial conformational changes in the enzyme upon nucleotide hydrolysis as well as a unique tetramer interface. Mutations of residues in this interface can affect tetramer formation, enzyme catalysis and regulation, indicating the functional importance of the tetramer. With altered glycolytic flux being a hallmark of cancers, these new structures allow a molecular understanding of the functional consequences of somatic PFK1 mutations identified in human cancers. We characterize three of these mutations and show they have distinct effects on allosteric regulation of PFKP activity and lactate production. The PFKP structural blueprint for somatic mutations as well as the catalytic site can guide therapeutic targeting of PFK1 activity to control dysregulated glycolysis in disease.
authors.affiliation: Department of Cell and Tissue Biology, University of California, San Francisco, California 94143, USA.
authors.forename: Bradley A
authors.initials: BA
authors.lastname: Webb
authors.affiliation: Department of Biological Sciences, Northeast Structural Genomics Consortium, Columbia University, New York, New York 10027, USA.
authors.forename: Farhad
authors.initials: F
authors.lastname: Forouhar
authors.affiliation: Department of Biological Sciences, Northeast Structural Genomics Consortium, Columbia University, New York, New York 10027, USA.
authors.forename: Fu-En
authors.initials: FE
authors.lastname: Szu
authors.affiliation: Department of Biological Sciences, Northeast Structural Genomics Consortium, Columbia University, New York, New York 10027, USA.
authors.forename: Jayaraman
authors.initials: J
authors.lastname: Seetharaman
authors.affiliation: Department of Biological Sciences, Northeast Structural Genomics Consortium, Columbia University, New York, New York 10027, USA.
authors.forename: Liang
authors.initials: L
authors.lastname: Tong
authors.affiliation: Department of Cell and Tissue Biology, University of California, San Francisco, California 94143, USA.
authors.forename: Diane L
authors.initials: DL
authors.lastname: Barber
mesh_terms: D004789:Enzyme Activation; D006801:Humans; D046529:Microscopy, Electron, Transmission; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D009369:Neoplasms / Q000201:enzymology* / Q000235:genetics; D010732:Phosphofructokinase-1 / Q000737:chemistry* / Q000235:genetics* / Q000648:ultrastructure; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D010732:Phosphofructokinase-1