Molecular cloning and sequencing of the human erythrocyte 2,3-bisphosphoglycerate mutase cDNA: revised amino acid sequence.

Abstract

The human erythrocyte 2,3‐bisphosphoglycerate mutase (BPGM) is a multifunctional enzyme which controls the metabolism of 2,3‐diphosphoglycerate, the main allosteric effector of haemoglobin. Several cDNA banks were constructed from reticulocyte mRNA, either by conventional cloning methods in pBR322 and screening with specific mixed oligonucleotide probes, or in the expression vector lambda gt 11. The largest cDNA isolated contained 1673 bases [plus the poly(A) tail], which is slightly smaller than the size of the intact mRNA as estimated by Northern blot analysis (approximately 1800 bases). This cDNA encodes for a protein of 258 residues; the protein yielded 34 tryptic peptides which were subsequently isolated by h.p.l.c. Our nucleotide sequence data were entirely confirmed by the amino acid composition of these tryptic peptides and reveal several major differences from the published sequence; the revised amino acid sequence of human BPGM is presented. These findings represent the first step in the study of the expression and regulation of this enzyme as a specific marker of the erythroid cell line.

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

“…In particular, His-179 is juxtaposed with the phosphorylated His-8 in the yeast PGM tertiary structure (11). This -170-amino acid spacing between active site histidines is closely reproduced in the related human muscle and brain PGM isozymes (26)(27)(28) and the human, rabbit, and mouse bisphosphoglycerate mutase enzymes (BPGM) (29)(30)(31).…”

“…The N-terminal His-8/258 (yeast PGM/rat Fru-2,6-P2ase) of the mutase/P2ase fold,.embedded in a conserved Arg-His-Gly-(Glu or Gln) motif, is matched by an analogous N-terminal histidine residue in a similar Arg-His-Gly-(Glu or Asp) acid RIF2,61PD 2 5 0 2 6 0 2 7 0 2 8 0 2 9 0 3 0 0 (26)(27)(28). BPGM sequences are from human (Hu) (29), rabbit (Rb) (30), and mouse (Mo) (31). The secondary structural elements of the yeast PGM crystal structure (11) are indicated below the alignment, 13-strands are labeled A-F, and a-helices are labeled 1-5.…”

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

“…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. In the first family, patients had severely reduced 2,3-DPG levels and increased ATP levels.…”

Recent progress in the molecular genetic analysis of erythroenzymopathy

“…There are two isozymes of MPGM: a muscle-specific form (MPGM-M) and non-muscle-specific form (MPGM-B) found in liver, kidney, brain, and red blood cells. The cDNAs for these enzymes have recently been cloned [ 19,20,24,25]. The DPGM cDNA encodes a protein of 258 amino acid residues [19,20].…”