Critical Role of Human Bisphosphoglycerate Mutase Cys22 in the Phosphatase Activator-binding Site

Ravel, Pascale; Craescu, Constantin T.; Arous, N.; Rosa, J.; Garel, Marie-Claude

doi:10.1074/jbc.272.22.14045

Cited by 10 publications

(9 citation statements)

References 40 publications

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“…Our ITC measurements suggest that citrate binds BPGAM with a K d value of 80 mM. The significance of this value is supported by the K m value of 300 mM for BPG (mutase activity of BPGAM; Ravel et al, 1997). Given that citrate and BPG share similar size and charge properties, they are likely to form similar multiple electrostatic interactions with the basic residues located at the entrance to the BPGAM active site.…”

Section: Discussionsupporting

confidence: 52%

Unliganded structure of human bisphosphoglycerate mutase reveals side-chain movements induced by ligand binding

2010

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PDB Reference: bisphosphoglycerate mutase, 3nfy.Erythrocyte-specific bisphosphoglycerate mutase is a trifunctional enzyme which modulates the levels of 2,3-bisphosphoglycerate (2,3-BPG) in red blood cells by virtue of its synthase and phosphatase activities. Low levels of erythrocyte 2,3-BPG increase the affinity of haemoglobin for oxygen, thus limiting the release of oxygen into tissues. 2,3-BPG levels in stored blood decline rapidly owing to the phosphatase activity of bisphosphoglycerate mutase, which is enhanced by a fall in pH. Here, the 1.94 Å resolution X-ray structure of bisphosphoglycerate mutase is presented, focusing on the dynamic nature of key ligand-binding residues and their interaction with the inhibitor citrate. Residues at the binding pocket are complete. In addition, the movement of key residues in the presence and absence of ligand is described and alternative conformations are explored. The conformation in which the ligand citrate would bind at the substrate-binding pocket is proposed, with discussion and representations of its orientation. The characterization of bisphosphoglycerate mutase-citrate interactions will provide a framework for the design of specific inhibitors of the phosphatase activity of this enzyme, which may limit the decline of 2,3-BPG in stored blood.

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Section: Discussionsupporting

confidence: 52%

Unliganded structure of human bisphosphoglycerate mutase reveals side-chain movements induced by ligand binding

2010

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“…The deletion of two or four residues gave partial loss of synthase activity. In addition, mutagenesis of Lys-245 implied its involvement at the active site [24]. These results would suggest that the role of the C-terminal region is to hold and to present Lys-245 in such a way as to contribute to ligand binding at the active site.…”

Section: Comparison With Bpgammentioning

confidence: 90%

The role of the C-terminal region in phosphoglycerate mutase

Walter

Nairn

Duncan

et al. 1998

Biochemical Journal

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Removal of the C-terminal seven residues from phosphoglycerate mutase from Saccharomyces cerevisiae by limited proteolysis is associated with loss of mutase activity, but no change in phosphatase activity. The presence of the cofactor 2, 3-bisphosphoglycerate, or of the cofactor and substrate 3-phosphoglycerate together, confers protection against proteolysis. The substrate alone offers no protection. Replacement of either or both of the two lysines at the C-terminus by glycines has only limited effects on the kinetic properties of phosphoglycerate mutase, indicating that these residues are unlikely to be involved in crucial electrostatic interactions with the substrate, intermediate or product in the reaction. However, the double-mutant form of the enzyme is more sensitive to proteolysis and is no longer protected against proteolysis by the presence of cofactor. The proteolysed wild-type and two of the mutated forms of the enzyme show a reduced response to 2-phosphoglycollate, which enhances the instability of the phospho form of the native enzyme. The phosphoglycerate mutase from Schizosaccharomyces pombe, which lacks the analogous C-terminal tail, has an inherently lower mutase activity and is also less responsive to stimulation by 2-phosphoglycollate. It is proposed that the C-terminal region of phosphoglycerate mutase helps to maintain the enzyme in its active phosphorylated form and assists in the retention of the bisphosphoglycerate intermediate at the active site. However, its role seems not to be to contribute directly to ligand binding, but rather to exert indirect effects on the transfer of the phospho group between substrate, enzyme, intermediate and product.

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“…The phosphatase reaction can be stimulated by a number of anions including chloride, phosphate, and particularly by 2-phosphoglycolate (5). These three enzymic activities have been found to occur at a unique active site with two different binding sites for the substrates, one for bisphosphoglycerate and another for monophosphoglycerate (6,7).…”

mentioning

confidence: 99%

Crystal Structure of Human Bisphosphoglycerate Mutase

Wang

Wei

Bian

et al. 2004

Journal of Biological Chemistry

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Bisphosphoglycerate mutase is a trifunctional enzyme of which the main function is to synthesize 2,3-bisphosphoglycerate, the allosteric effector of hemoglobin. The gene coding for bisphosphoglycerate mutase from the human cDNA library was cloned and expressed in Escherichia coli. The protein crystals were obtained and diffract to 2.5 Å and produced the first crystal structure of bisphosphoglycerate mutase. The model was refined to a crystallographic R-factor of 0.200 and R free of 0.266 with excellent stereochemistry. The enzyme remains a dimer in the crystal. The overall structure of the enzyme resembles that of the cofactor-dependent phosphoglycerate mutase except the regions of 13-21, 98 -117, 127-151, and the C-terminal tail. The conformational changes in the backbone and the side chains of some residues reveal the structural basis for the different activities between phosphoglycerate mutase and bisphosphoglycerate mutase. The bisphosphoglycerate mutase-specific residue Gly-14 may cause the most important conformational changes, which makes the side chain of Glu-13 orient toward the active site. The positions of Glu-13 and Phe-22 prevent 2,3-bisphosphoglycerate from binding in the way proposed previously. In addition, the side chain of Glu-13 would affect the Glu-89 protonation ability responsible for the low mutase activity. Other structural variations, which could be connected with functional differences, are also discussed.Bisphosphoglycerate mutase (BPGM) 1 is an erythrocyte-specific trifunctional enzyme. The main activity is that of synthase (BPGM, EC 5.4.2.4), catalyzing the formation of 2,3-bisphosphoglycerate (2,3-BPG) from 1,3-bisphosphoglycerate (1,3-BPG) (1-3) (see Fig. 1A). The second activity is that of a mutase (phosphoglycerate mutase, EC 5.4.2.1) catalyzing the interconversion between 2-and 3-phosphoglycerate (Fig. 1B) (4). The third activity, as a phosphatase (S-succinylglutathione hydrolase, EC 3.1.3.13), is to catalyze the hydrolysis of 2,3-BPG to 3-or 2-phosphoglycerate and a phosphate (Fig. 1C) (1, 2). The phosphatase reaction can be stimulated by a number of anions including chloride, phosphate, and particularly by 2-phosphoglycolate (5). These three enzymic activities have been found to occur at a unique active site with two different binding sites for the substrates, one for bisphosphoglycerate and another for monophosphoglycerate (6, 7).BPGM regulates the level of 2,3-BPG in human blood cells. In vivo, the concentration of 2,3-BPG is determined by the relative activities of the synthase and phosphatase reactions. In red blood cells, 2,3-BPG is the main allosteric effector of hemoglobin. It shifts the equilibrium between the oxy and deoxy conformations of hemoglobins by preferentially stabilizing the unliganded form. Sickle cell anemia is characterized by polymerization of deoxygenated hemoglobin mutants giving rise to deformed erythrocytes and vaso-occlusive complications. 2,3-BPG has been shown to facilitate this polymerization. The ability to modulate the 2,3-BPG level in vivo wou...

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Critical Role of Human Bisphosphoglycerate Mutase Cys22 in the Phosphatase Activator-binding Site

Cited by 10 publications

References 40 publications

Unliganded structure of human bisphosphoglycerate mutase reveals side-chain movements induced by ligand binding

Unliganded structure of human bisphosphoglycerate mutase reveals side-chain movements induced by ligand binding

The role of the C-terminal region in phosphoglycerate mutase

Crystal Structure of Human Bisphosphoglycerate Mutase

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