Crystal Structure of Human Bisphosphoglycerate Mutase

Abstract

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...

Influence of torpor on cardiac expression of genes involved in the circadian clock and protein turnover in the Siberian hamster (Phodopus sungorus)

“…The globins might act similarly in Siberian hamsters, although because all globin transcripts are suppressed during torpor and arousal, they would be implicated in winter normothermia rather than torpor. BPGM catalyzes the synthesis of 2,3-BPG, which decreases the affinity of hemoglobin for oxygen (42). Therefore, downregulation of this transcript may help to increase oxygen transport during torpor, possibly preventing hypoxia due to decreased respiration rates during torpor.…”

Seeing the Process of Histidine Phosphorylation in Human Bisphosphoglycerate Mutase

“…The expression and purification of human BPGM were performed as described previously (8). All crystals were grown using the hanging drop vapor diffusion method.…”

“…The quality and stereochemistry of the final models were analyzed with PROCHECK (23). In the Ramachandran plots of all structures, more than 91.8% of the residues are located in the most-favored regions, and only Ser-24 is located in the general allowed region as in the free BPGM structure (8). The statistics of data collection and structure refinement are listed in Table 1.…”

“…Both BPGM and dPGM can catalyze the same three overall reactions for which phosphorylation of the catalytic histidine by 2,3-bisphosphoglycerate is the first step. Previous studies of BPGM and dPGMs have provided important but so far incomplete information on the structural basis for the specific activities and molecular mechanisms (5)(6)(7)(8)(9)(10)(11)(12)(13)(14). Although a number of structures of dPGMs (9,10,12,13,(15)(16)(17)(18) and human BPGM (8) have been reported, no structure of the enzyme⅐substrate complex is available.…”

The energy-less red blood cell is lost: erythrocyte enzyme abnormalities of glycolysis

“…108 Recently, the crystal structure of human BPGM has been determined (PDB entry 1T8P), confirming the concept that BPGM and MPGM are structurally homologous enzymes, and providing a rationale for the specific residues that are crucial for synthase, mutase, and phosphatase activity. 109 The gene for BPGM (BPGM) has been mapped to chromosome 7q31-34 and it consists of 3 exons, spanning more than 22 kb. 110 BPGM deficiency (OMIM 222 800) is a very rare autosomal recessive disorder and only 2 affected families have been described.…”