ID F264_RAT Reviewed; 469 AA. AC P25114; DT 01-MAY-1992, integrated into UniProtKB/Swiss-Prot. DT 23-JAN-2007, sequence version 3. DT 27-MAR-2024, entry version 179. DE RecName: Full=6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 4; DE Short=6PF-2-K/Fru-2,6-P2ase 4; DE Short=PFK/FBPase 4; DE AltName: Full=6PF-2-K/Fru-2,6-P2ase testis-type isozyme; DE Includes: DE RecName: Full=6-phosphofructo-2-kinase; DE EC=2.7.1.105 {ECO:0000269|PubMed:1651918}; DE Includes: DE RecName: Full=Fructose-2,6-bisphosphatase; DE EC=3.1.3.46 {ECO:0000269|PubMed:1651918}; GN Name=Pfkfb4; OS Rattus norvegicus (Rat). OC Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi; Mammalia; OC Eutheria; Euarchontoglires; Glires; Rodentia; Myomorpha; Muroidea; Muridae; OC Murinae; Rattus. OX NCBI_TaxID=10116; RN [1] RP NUCLEOTIDE SEQUENCE [MRNA], TISSUE SPECIFICITY, FUNCTION, CATALYTIC RP ACTIVITY, BIOPHYSICOCHEMICAL PROPERTIES, SUBUNIT, AND ACTIVITY REGULATION. RC TISSUE=Testis; RX PubMed=1651918; DOI=10.1016/s0021-9258(18)98475-9; RA Sakata J., Abe Y., Uyeda K.; RT "Molecular cloning of the DNA and expression and characterization of rat RT testes fructose-6-phosphate,2-kinase:fructose-2,6-bisphosphatase."; RL J. Biol. Chem. 266:15764-15770(1991). RN [2] RP X-RAY CRYSTALLOGRAPHY (2.0 ANGSTROMS) IN COMPLEX WITH ATP ANALOG. RC TISSUE=Testis; RX PubMed=8805587; DOI=10.1016/s0969-2126(96)00109-8; RA Hasemann C.A., Istvan E.S., Uyeda K., Deisenhofer J.; RT "The crystal structure of the bifunctional enzyme 6-phosphofructo-2- RT kinase/fructose-2,6-bisphosphatase reveals distinct domain homologies."; RL Structure 4:1017-1029(1996). RN [3] RP X-RAY CRYSTALLOGRAPHY (2.4 ANGSTROMS) OF MUTANT ALA-257 IN COMPLEX WITH ATP RP ANALOG; FRUCTOSE 6-PHOSPHATE AND PHOSPHATE ION, SUBUNIT, AND ACTIVE SITE. RC TISSUE=Testis; RX PubMed=9890980; DOI=10.1074/jbc.274.4.2176; RA Yuen M.H., Mizuguchi H., Lee Y.H., Cook P.F., Uyeda K., Hasemann C.A.; RT "Crystal structure of the H256A mutant of rat testis fructose-6- RT phosphate,2-kinase/fructose-2,6-bisphosphatase. Fructose 6-phosphate in the RT active site leads to mechanisms for both mutant and wild type RT bisphosphatase activities."; RL J. Biol. Chem. 274:2176-2184(1999). CC -!- FUNCTION: Synthesis and degradation of fructose 2,6-bisphosphate. CC {ECO:0000269|PubMed:1651918}. CC -!- CATALYTIC ACTIVITY: CC Reaction=beta-D-fructose 2,6-bisphosphate + H2O = beta-D-fructose 6- CC phosphate + phosphate; Xref=Rhea:RHEA:17289, ChEBI:CHEBI:15377, CC ChEBI:CHEBI:43474, ChEBI:CHEBI:57634, ChEBI:CHEBI:58579; EC=3.1.3.46; CC Evidence={ECO:0000269|PubMed:1651918}; CC PhysiologicalDirection=left-to-right; Xref=Rhea:RHEA:17290; CC Evidence={ECO:0000305|PubMed:1651918}; CC -!- CATALYTIC ACTIVITY: CC Reaction=ATP + beta-D-fructose 6-phosphate = ADP + beta-D-fructose 2,6- CC bisphosphate + H(+); Xref=Rhea:RHEA:15653, ChEBI:CHEBI:15378, CC ChEBI:CHEBI:30616, ChEBI:CHEBI:57634, ChEBI:CHEBI:58579, CC ChEBI:CHEBI:456216; EC=2.7.1.105; CC Evidence={ECO:0000269|PubMed:1651918}; CC PhysiologicalDirection=left-to-right; Xref=Rhea:RHEA:15654; CC Evidence={ECO:0000305|PubMed:1651918}; CC -!- ACTIVITY REGULATION: The most important regulatory mechanism of these CC opposing activities is by phosphorylation and dephosphorylation of the CC enzyme. {ECO:0000269|PubMed:1651918}. CC -!- BIOPHYSICOCHEMICAL PROPERTIES: CC Kinetic parameters: CC KM=85 uM for beta-D-fructose 6-phosphate CC {ECO:0000269|PubMed:1651918}; CC KM=270 uM for ATP {ECO:0000269|PubMed:1651918}; CC KM=21 uM for beta-D-fructose 2,6-bisphosphate CC {ECO:0000269|PubMed:1651918}; CC -!- SUBUNIT: Homodimer. {ECO:0000269|PubMed:1651918, CC ECO:0000269|PubMed:8805587, ECO:0000269|PubMed:9890980}. CC -!- TISSUE SPECIFICITY: Testis. {ECO:0000269|PubMed:1651918}. CC -!- SIMILARITY: In the C-terminal section; belongs to the phosphoglycerate CC mutase family. {ECO:0000305}. CC --------------------------------------------------------------------------- CC Copyrighted by the UniProt Consortium, see https://www.uniprot.org/terms CC Distributed under the Creative Commons Attribution (CC BY 4.0) License CC --------------------------------------------------------------------------- DR EMBL; M64797; AAA41163.1; -; mRNA. DR PIR; A40800; A40800. DR RefSeq; NP_062206.1; NM_019333.1. DR RefSeq; XP_008764785.1; XM_008766563.2. DR RefSeq; XP_017459205.1; XM_017603716.1. DR PDB; 1BIF; X-ray; 2.00 A; A=1-469. DR PDB; 2BIF; X-ray; 2.40 A; A/B=1-469. DR PDB; 3BIF; X-ray; 2.30 A; A=2-469. DR PDBsum; 1BIF; -. DR PDBsum; 2BIF; -. DR PDBsum; 3BIF; -. DR AlphaFoldDB; P25114; -. DR SMR; P25114; -. DR ComplexPortal; CPX-2045; 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 4 complex. DR DIP; DIP-2904N; -. DR STRING; 10116.ENSRNOP00000074158; -. DR PhosphoSitePlus; P25114; -. DR jPOST; P25114; -. DR PaxDb; 10116-ENSRNOP00000028039; -. DR Ensembl; ENSRNOT00000028039.5; ENSRNOP00000028039.2; ENSRNOG00000020656.7. DR Ensembl; ENSRNOT00055011984; ENSRNOP00055009488; ENSRNOG00055007209. DR Ensembl; ENSRNOT00060030716; ENSRNOP00060024865; ENSRNOG00060017864. DR Ensembl; ENSRNOT00065010840; ENSRNOP00065007992; ENSRNOG00065006921. DR GeneID; 54283; -. DR KEGG; rno:54283; -. DR UCSC; RGD:3310; rat. DR AGR; RGD:3310; -. DR CTD; 5210; -. DR RGD; 3310; Pfkfb4. DR eggNOG; KOG0234; Eukaryota. DR GeneTree; ENSGT00950000182835; -. DR HOGENOM; CLU_006383_1_1_1; -. DR InParanoid; P25114; -. DR OMA; RCLMGYF; -. DR OrthoDB; 2013830at2759; -. DR PhylomeDB; P25114; -. DR TreeFam; TF313541; -. DR BRENDA; 2.7.1.105; 5301. DR BRENDA; 3.1.3.46; 5301. DR Reactome; R-RNO-9634600; Regulation of glycolysis by fructose 2,6-bisphosphate metabolism. DR SABIO-RK; P25114; -. DR EvolutionaryTrace; P25114; -. DR PRO; PR:P25114; -. DR Proteomes; UP000002494; Chromosome 8. DR Bgee; ENSRNOG00000020656; Expressed in jejunum and 18 other cell types or tissues. DR ExpressionAtlas; P25114; baseline. DR GO; GO:0005829; C:cytosol; IBA:GO_Central. DR GO; GO:0003873; F:6-phosphofructo-2-kinase activity; IDA:RGD. DR GO; GO:0005524; F:ATP binding; IEA:UniProtKB-KW. DR GO; GO:0004331; F:fructose-2,6-bisphosphate 2-phosphatase activity; IDA:RGD. DR GO; GO:0006003; P:fructose 2,6-bisphosphate metabolic process; IBA:GO_Central. DR GO; GO:0006000; P:fructose metabolic process; IEA:InterPro. DR CDD; cd07067; HP_PGM_like; 1. DR Gene3D; 3.40.50.300; P-loop containing nucleotide triphosphate hydrolases; 1. DR Gene3D; 3.40.50.1240; Phosphoglycerate mutase-like; 1. DR InterPro; IPR003094; 6Pfruct_kin. DR InterPro; IPR013079; 6Phosfructo_kin. DR InterPro; IPR013078; His_Pase_superF_clade-1. DR InterPro; IPR029033; His_PPase_superfam. DR InterPro; IPR027417; P-loop_NTPase. DR InterPro; IPR001345; PG/BPGM_mutase_AS. DR PANTHER; PTHR10606; 6-PHOSPHOFRUCTO-2-KINASE/FRUCTOSE-2,6-BISPHOSPHATASE; 1. DR PANTHER; PTHR10606:SF14; 6-PHOSPHOFRUCTO-2-KINASE_FRUCTOSE-2,6-BISPHOSPHATASE 4; 1. DR Pfam; PF01591; 6PF2K; 1. DR Pfam; PF00300; His_Phos_1; 1. DR PIRSF; PIRSF000709; 6PFK_2-Ptase; 1. DR PRINTS; PR00991; 6PFRUCTKNASE. DR SMART; SM00855; PGAM; 1. DR SUPFAM; SSF52540; P-loop containing nucleoside triphosphate hydrolases; 1. DR SUPFAM; SSF53254; Phosphoglycerate mutase-like; 1. DR PROSITE; PS00175; PG_MUTASE; 1. DR Genevisible; P25114; RN. PE 1: Evidence at protein level; KW 3D-structure; ATP-binding; Hydrolase; Kinase; Multifunctional enzyme; KW Nucleotide-binding; Phosphoprotein; Reference proteome; Transferase. FT CHAIN 1..469 FT /note="6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase FT 4" FT /id="PRO_0000179971" FT REGION 1..249 FT /note="6-phosphofructo-2-kinase" FT /evidence="ECO:0000305|PubMed:1651918" FT REGION 250..469 FT /note="Fructose-2,6-bisphosphatase" FT /evidence="ECO:0000305|PubMed:1651918" FT ACT_SITE 129 FT /evidence="ECO:0000255" FT ACT_SITE 159 FT /evidence="ECO:0000255" FT ACT_SITE 257 FT /note="Tele-phosphohistidine intermediate" FT /evidence="ECO:0000269|PubMed:9890980" FT ACT_SITE 326 FT /note="Proton donor/acceptor" FT /evidence="ECO:0000269|PubMed:9890980" FT BINDING 46..54 FT /ligand="ATP" FT /ligand_id="ChEBI:CHEBI:30616" FT /evidence="ECO:0000269|PubMed:8805587" FT BINDING 79 FT /ligand="beta-D-fructose 6-phosphate" FT /ligand_id="ChEBI:CHEBI:57634" FT /evidence="ECO:0000269|PubMed:9890980" FT BINDING 103 FT /ligand="beta-D-fructose 6-phosphate" FT /ligand_id="ChEBI:CHEBI:57634" FT /evidence="ECO:0000250|UniProtKB:Q16875" FT BINDING 131 FT /ligand="beta-D-fructose 6-phosphate" FT /ligand_id="ChEBI:CHEBI:57634" FT /evidence="ECO:0000269|PubMed:9890980" FT BINDING 137 FT /ligand="beta-D-fructose 6-phosphate" FT /ligand_id="ChEBI:CHEBI:57634" FT /evidence="ECO:0000269|PubMed:9890980" FT BINDING 168..173 FT /ligand="ATP" FT /ligand_id="ChEBI:CHEBI:30616" FT /evidence="ECO:0000269|PubMed:8805587" FT BINDING 173 FT /ligand="beta-D-fructose 6-phosphate" FT /ligand_id="ChEBI:CHEBI:57634" FT /evidence="ECO:0000250|UniProtKB:Q16875" FT BINDING 194 FT /ligand="beta-D-fructose 6-phosphate" FT /ligand_id="ChEBI:CHEBI:57634" FT /evidence="ECO:0000250|UniProtKB:Q16875" FT BINDING 198 FT /ligand="beta-D-fructose 6-phosphate" FT /ligand_id="ChEBI:CHEBI:57634" FT /evidence="ECO:0000269|PubMed:9890980" FT BINDING 256 FT /ligand="beta-D-fructose 2,6-bisphosphate" FT /ligand_id="ChEBI:CHEBI:58579" FT /evidence="ECO:0000250|UniProtKB:Q16875" FT BINDING 263 FT /ligand="beta-D-fructose 2,6-bisphosphate" FT /ligand_id="ChEBI:CHEBI:58579" FT /evidence="ECO:0000250|UniProtKB:Q16875" FT BINDING 269 FT /ligand="beta-D-fructose 2,6-bisphosphate" FT /ligand_id="ChEBI:CHEBI:58579" FT /evidence="ECO:0000269|PubMed:9890980" FT BINDING 306 FT /ligand="beta-D-fructose 2,6-bisphosphate" FT /ligand_id="ChEBI:CHEBI:58579" FT /evidence="ECO:0000250|UniProtKB:Q16875" FT BINDING 337 FT /ligand="beta-D-fructose 2,6-bisphosphate" FT /ligand_id="ChEBI:CHEBI:58579" FT /evidence="ECO:0000269|PubMed:9890980" FT BINDING 348..351 FT /ligand="ATP" FT /ligand_id="ChEBI:CHEBI:30616" FT /evidence="ECO:0000250|UniProtKB:P07953" FT BINDING 351 FT /ligand="beta-D-fructose 2,6-bisphosphate" FT /ligand_id="ChEBI:CHEBI:58579" FT /evidence="ECO:0000269|PubMed:9890980" FT BINDING 355 FT /ligand="beta-D-fructose 2,6-bisphosphate" FT /ligand_id="ChEBI:CHEBI:58579" FT /evidence="ECO:0000269|PubMed:9890980" FT BINDING 366 FT /ligand="beta-D-fructose 2,6-bisphosphate" FT /ligand_id="ChEBI:CHEBI:58579" FT /evidence="ECO:0000269|PubMed:9890980" FT BINDING 392..396 FT /ligand="ATP" FT /ligand_id="ChEBI:CHEBI:30616" FT /evidence="ECO:0000250|UniProtKB:P07953" FT BINDING 392 FT /ligand="beta-D-fructose 2,6-bisphosphate" FT /ligand_id="ChEBI:CHEBI:58579" FT /evidence="ECO:0000269|PubMed:9890980" FT BINDING 396 FT /ligand="beta-D-fructose 2,6-bisphosphate" FT /ligand_id="ChEBI:CHEBI:58579" FT /evidence="ECO:0000269|PubMed:9890980" FT BINDING 428 FT /ligand="ATP" FT /ligand_id="ChEBI:CHEBI:30616" FT /evidence="ECO:0000269|PubMed:8805587" FT SITE 391 FT /note="Transition state stabilizer" FT /evidence="ECO:0000250|UniProtKB:P00950" FT MOD_RES 29 FT /note="Phosphoserine; by PKC" FT /evidence="ECO:0000255" FT MOD_RES 444 FT /note="Phosphothreonine; by PKC" FT /evidence="ECO:0000255" FT STRAND 40..45 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 52..65 FT /evidence="ECO:0007829|PDB:1BIF" FT STRAND 70..74 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 75..83 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 89..92 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 97..119 FT /evidence="ECO:0007829|PDB:1BIF" FT STRAND 124..130 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 135..148 FT /evidence="ECO:0007829|PDB:1BIF" FT STRAND 151..157 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 162..172 FT /evidence="ECO:0007829|PDB:1BIF" FT TURN 173..175 FT /evidence="ECO:0007829|PDB:1BIF" FT TURN 177..181 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 184..199 FT /evidence="ECO:0007829|PDB:1BIF" FT TURN 207..212 FT /evidence="ECO:0007829|PDB:1BIF" FT STRAND 215..218 FT /evidence="ECO:0007829|PDB:1BIF" FT TURN 220..222 FT /evidence="ECO:0007829|PDB:1BIF" FT STRAND 225..228 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 233..242 FT /evidence="ECO:0007829|PDB:1BIF" FT STRAND 252..256 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 261..265 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 276..292 FT /evidence="ECO:0007829|PDB:1BIF" FT STRAND 298..301 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 305..311 FT /evidence="ECO:0007829|PDB:1BIF" FT STRAND 314..316 FT /evidence="ECO:0007829|PDB:1BIF" FT STRAND 318..320 FT /evidence="ECO:0007829|PDB:3BIF" FT HELIX 322..324 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 330..332 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 337..343 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 345..353 FT /evidence="ECO:0007829|PDB:1BIF" FT TURN 355..357 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 366..382 FT /evidence="ECO:0007829|PDB:1BIF" FT STRAND 384..390 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 392..402 FT /evidence="ECO:0007829|PDB:1BIF" FT TURN 407..409 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 410..412 FT /evidence="ECO:0007829|PDB:1BIF" FT STRAND 419..425 FT /evidence="ECO:0007829|PDB:1BIF" FT STRAND 427..436 FT /evidence="ECO:0007829|PDB:1BIF" FT HELIX 458..461 FT /evidence="ECO:0007829|PDB:1BIF" FT TURN 462..464 FT /evidence="ECO:0007829|PDB:2BIF" SQ SEQUENCE 469 AA; 54155 MW; E3D0AF34D3A09CA6 CRC64; MASPRELTQN PLKKIWMPYS NGRPALHASQ RGVCMTNCPT LIVMVGLPAR GKTYISKKLT RYLNWIGVPT REFNVGQYRR DMVKTYKSFE FFLPDNEEGL KIRKQCALAA LNDVRKFLSE EGGHVAVFDA TNTTRERRAM IFNFGEQNGY KTFFVESICV DPEVIAANIV QVKLGSPDYV NRDSDEATED FMRRIECYEN SYESLDEEQD RDLSYIKIMD VGQSYVVNRV ADHIQSRIVY YLMNIHVTPR SIYLCRHGES ELNLKGRIGG DPGLSPRGRE FSKHLAQFIS DQNIKDLKVW TSQMKRTIQT AEALSVPYEQ WKVLNEIDAG VCEEMTYEEI QDHYPLEFAL RDQDKYRYRY PKGESYEDLV QRLEPVIMEL ERQENVLVIC HQAVMRCLLA YFLDKAAEEL PYLKCPLHTV LKLTPVAYGC KVESIFLNVA AVNTHRDRPQ NVDISRPSEE ALVTVPAHQ //