Evolution of a bifunctional enzyme: 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase.

Bazán, J. Fernando; Fletterick, R.J.; Pilkis, S J

doi:10.1073/pnas.86.24.9642

Cited by 101 publications

(71 citation statements)

References 49 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Alignment of amino acid sequences of acid phosphatases (Roiko et al, 1990;Van Etten et al, 1991) has revealed a conserved sequence motif RHGXRXP. The RHG sequence is also found in phosphoglycerate mutase and 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase which suggested that these three enzymes might share a common fold (Bazan et al, 1989). Recent crystallographic studies Lindqvist et al, 1993) of native rat prostatic acid phosphatase and its complex with the inhibitor L(+)-tartrate have shown that the structure of acid phosphatase consists of two domains, an utP domain consisting of a seven-stranded mixed P-sheet with a-helices on both sides of the sheet, and a smaller ahelical domain.…”

mentioning

confidence: 98%

Crystal structures of rat acid phosphatase complexed with the transition‐state analogs vanadate and molybdate

Lindqvist

Schneider

Vihko

1994

European Journal of Biochemistry

132

125

View full text Add to dashboard Cite

The three-dimensional structures of complexes of recombinant rat prostatic acid phosphatase with the transition-state analogs vanadate and molybdate were determined to 0.3-nm resolution using protein crystallographic methods. The overall structure of the enzyme remains unchanged upon binding of the metal oxyanions; only local conformational differences in the positions of some side chains at the active site were found. The metal oxyanions bind in an identical fashion at the active site with trigonal bipyramidal coordination geometry. The metal ion is within coordination distance of the His12 side chain which is located at one of the axial positions. The three equatorial oxygen atoms interact with the conserved residues Argll, Argl.5, Arg79 and His257. Within hydrogen-bonding distance of the axial oxygen atom is the side chain of the conserved residue Asp258. The implications of these results for the catalytic mechanism of acid phosphatase are discussed.The hydrolysis of phosphoric monoesters is catalyzed by a number of different, non-related enzymes. High-molecularmass acid phosphatases (ortho-phosphoric-monoester phosphohydrolases) form one class of phosphoric-monoester phosphatases which also catalyze phosphoryl transfer between a phosphoester and alcohols (Bodansky, 1972;Van Etten, 1982;Vincent et al., 1992). Alignment of amino acid sequences of acid phosphatases (Roiko et al., 1990;Van Etten et al., 1991) has revealed a conserved sequence motif RHGXRXP. The RHG sequence is also found in phosphoglycerate mutase and 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase which suggested that these three enzymes might share a common fold (Bazan et al., 1989). Recent crystallographic studies Lindqvist et al., 1993) of native rat prostatic acid phosphatase and its complex with the inhibitor L(+)-tartrate have shown that the structure of acid phosphatase consists of two domains, an utP domain consisting of a seven-stranded mixed P-sheet with a-helices on both sides of the sheet, and a smaller ahelical domain. The overall fold of the alp domain is indeed similar to the fold observed in phosphoglycerate mutase. The active site is located at the C-terminus of the P-strands in the alp domain and the conserved residues Argll, Hisl2, Argl5, Arg79 are part of the phosphate-binding site. In the binary complex with the inhibitor L( +)-tartrate, these residues and the conserved residues His257 and Asp258 are involved in binding of the inhibitor. Site-directed mutagenesis studies of acid phosphatase from Escherichia coli showed that the residues corresponding to Argll and His12 are required for catalysis and that substitutions of residues corresponding to Arg15, Arg79, His257 and Asp258 severely impair the catalytic activity (Ostanin et al., 1992;Ostanin and Van Etten, 1993) consistent with the location of these residues at the active site. The chemical pathway of phosphoric-ester hydrolysis by acid phosphatases is summarized in Scheme 1 and is supported by many studies (Van Etten, 1982;Vincent et al., 1992). The hydrolysis can...

show abstract

mentioning

confidence: 98%

Crystal structures of rat acid phosphatase complexed with the transition‐state analogs vanadate and molybdate

Lindqvist

Schneider

Vihko

1994

European Journal of Biochemistry

132

125

View full text Add to dashboard Cite

show abstract

“…Because this enzyme controls the cellular concentration of fructose-2,6-bisphosphate (Fru-2,6-P 2 ), which is the most potent allosteric activator of phosphofructokinase, the rate-limiting enzyme of glycolysis, PFKFB eventually controls the glycolytic rate (1)(2)(3). The bifunctional enzyme acutely controls the concentration of Fru-2,6-P 2 by modulating the two mutually exclusive catalytic activities of Fru-2,6-P 2 synthesis (2-Kase) and hydrolysis (2-Pase) that reside in the two separate domains (4,5). The two activities are exquisitely regulated by various metabolic products and signal transduction-dependent phosphorylation such that the resulting predominant activity determines the final concentration of Fru-2,6-P 2 and, ultimately, the rates of glycolysis (6 -9).…”

mentioning

confidence: 99%

Crystal Structure of the Hypoxia-inducible Form of 6-Phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFKFB3)

Kim

Manes

el-Maghrabi

et al. 2006

Journal of Biological Chemistry

View full text Add to dashboard Cite

The hypoxia-inducible form of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFKFB3) plays a crucial role in the progression of cancerous cells by enabling their glycolytic pathways even under severe hypoxic conditions. To understand its structural architecture and to provide a molecular scaffold for the design of new cancer therapeutics, the crystal structure of the human form was determined. The structure at 2.1 Å resolution shows that the overall folding and functional dimerization are very similar to those of the liver (PFKFB1) and testis (PFKFB4) forms, as expected from sequence homology. However, in this structure, the N-terminal regulatory domain is revealed for the first time among the PFKFB isoforms. With a ␤-hairpin structure, the N terminus interacts with the 2-Pase domain to secure binding of fructose-6-phosphate to the active pocket, slowing down the release of fructose-6-phosphate from the phosphoenzyme intermediate product complex. The C-terminal regulatory domain is mostly disordered, leaving the active pocket of the fructose-2,6-bisphosphatase domain wide open. The active pocket of the 6-phosphofructo-2-kinase domain has a more rigid conformation, allowing independent bindings of substrates, fructose-6-phosphate and ATP, with higher affinities than other isoforms. Intriguingly, the structure shows an EDTA molecule bound to the fructose-6-phosphate site of the 6-phosphofructo-2-kinase active pocket despite its unfavorable liganding concentration, suggesting a high affinity. EDTA is not removable from the site with fructose-6-P alone but is with both ATP and fructose-6-P or with fructose-2,6-bisphosphate. This finding suggests that a molecule in which EDTA is covalently linked to ADP is a good starting molecule for the development of new cancer-therapeutic molecules.

show abstract

“…Shaded boxes contain conserved catalytic and substrate binding sites [12]. Using numbering based on rat liver sequence, these sites include: Arg452, Lys-356, and Arg-360, residues shown to bind substrate and/or product which are found on a surface loop of all mammalian FBPases; His-258, Glu-327 and His-392, a trio of catalytic residues in rat liver [30,12]; Arg-195, a key residue involved in Fru-6-P binding [31]; Arg-257 and Arg-307, which associate with the reactive C-2 phospho group of Fru-2,6-P2 [30,32]; and the ATP binding site, GlyLeu-Pro-Ala-Arg-Gly-Lys-Thr [33]. Underlined residues represent potential phosphorylation sites.…”

Section: Discussionmentioning

confidence: 99%

Molecular cloning and tissue‐specific expression of mouse kidney 6‐phosphofructo‐2‐kinase/fructose‐2,6‐bisphosphatase

Batra

Brown

et al. 1996

FEBS Letters

View full text Add to dashboard Cite

A 1932 bp cDNA clone encoding a novel isozyme of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFK-2/ FBPase-2) was isolated from a mouse kidney cDNA library. The sequence encodes 519 amino acids and, based on homology to rat heart genomic sequence, appears to be the product of alternative splicing from PFK-2/FBPase-2 gene B with an extended version of exon 15. Northern blot analysis indicated that this clone corresponds to an 8 kb mRNA expressed in multiple tissues, with the strongest signal in kidney, and detects several additional transcripts which may be alternatively spliced from gene B.

show abstract

Evolution of a bifunctional enzyme: 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase.

Abstract: The bifunctional rat liver enzyme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (ATP:Dfructose-6-phosphate 2-phosphotransferase/D-fructose-2,6-bisphosphate 2-phosphohydrolase, EC 2.7

Cited by 101 publications

References 49 publications

Crystal structures of rat acid phosphatase complexed with the transition‐state analogs vanadate and molybdate

Crystal structures of rat acid phosphatase complexed with the transition‐state analogs vanadate and molybdate

Crystal Structure of the Hypoxia-inducible Form of 6-Phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFKFB3)

Molecular cloning and tissue‐specific expression of mouse kidney 6‐phosphofructo‐2‐kinase/fructose‐2,6‐bisphosphatase

Contact Info

Product

Resources

About