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. 1999 Nov;8(11):2438–2443. doi: 10.1110/ps.8.11.2438

Probing the location and function of the conserved histidine residue of phosphoglucose isomerase by using an active site directed inhibitor N-bromoacetylethanolamine phosphate.

M Meng 1, T L Chane 1, Y J Sun 1, C D Hsiao 1
PMCID: PMC2144184  PMID: 10595547

Abstract

Phosphoglucose isomerase (EC 5.3.1.9) catalyzes the interconversion of D-glucopyranose-6-phosphate and D-fructofuranose-6-phosphate by promoting an intrahydrogen transfer between C1 and C2. A conserved histidine exists throughout all phosphoglucose isomerases and was hypothesized to be the base catalyzing the isomerization reaction. In the present study, this conserved histidine, His311, of the enzyme from Bacillus stearothermophilus was subjected to mutational analysis, and the mutational effect on the inactivation kinetics by N-bromoacetylethanolamine phosphate was investigated. The substitution of His311 with alanine, asparagine, or glutamine resulted in the decrease of activity, in k(cat)/K(M), by a factor of 10(3), indicating the importance of this residue. N-bromoacetylethanolamine phosphate inactivated irreversibly the activity of wild-type phosphoglucose isomerase; however, His311 --> Ala became resistant to this inhibitor, indicating that His311 is located in the active site and is responsible for the inactivation of the enzyme by this active site-directed inhibitor. The pKa of His311 was estimated to be 6.31 according to the pH dependence of the inactivation. The proximity of this value with the pKa value of 6.35, determined from the pH dependence of k(cat)/K(M), supports a role of His311 as a general base in the catalysis.

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Selected References

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  1. Beutler E., West C., Britton H. A., Harris J., Forman L. Glucosephosphate isomerase (GPI) deficiency mutations associated with hereditary nonspherocytic hemolytic anemia (HNSHA). Blood Cells Mol Dis. 1997 Dec;23(3):402–409. doi: 10.1006/bcmd.1997.0157. [DOI] [PubMed] [Google Scholar]
  2. Chaput M., Claes V., Portetelle D., Cludts I., Cravador A., Burny A., Gras H., Tartar A. The neurotrophic factor neuroleukin is 90% homologous with phosphohexose isomerase. Nature. 1988 Mar 31;332(6163):454–455. doi: 10.1038/332454a0. [DOI] [PubMed] [Google Scholar]
  3. Dyson J. E., Noltmann E. A. The effect of pH and temperature on the kinetic parameters of phosphoglucose isomerase. Participation of histidine and lysine in a proposed dual function mechanism. J Biol Chem. 1968 Apr 10;243(7):1401–1414. [PubMed] [Google Scholar]
  4. Faik P., Walker J. I., Redmill A. A., Morgan M. J. Mouse glucose-6-phosphate isomerase and neuroleukin have identical 3' sequences. Nature. 1988 Mar 31;332(6163):455–457. doi: 10.1038/332455a0. [DOI] [PubMed] [Google Scholar]
  5. Froman B. E., Tait R. C., Gottlieb L. D. Isolation and characterization of the phosphoglucose isomerase gene from Escherichia coli. Mol Gen Genet. 1989 May;217(1):126–131. doi: 10.1007/BF00330951. [DOI] [PubMed] [Google Scholar]
  6. Gibson D. R., Gracy R. W., Hartman F. C. Affinity labeling and characterization of the active site histidine of glucosephosphate isomerase. Sequence homology with triosephosphate isomerase. J Biol Chem. 1980 Oct 10;255(19):9369–9374. [PubMed] [Google Scholar]
  7. Hartman F. C., Suh B., Welch M. H., Barker R. Inactivation of class I fructose diphosphate aldolases by the substrate analog N-bromoacetylethanolamine phosphate. J Biol Chem. 1973 Dec 10;248(23):8233–8239. [PubMed] [Google Scholar]
  8. Hesman T. L., Barnell W. O., Conway T. Cloning, characterization, and nucleotide sequence analysis of a Zymomonas mobilis phosphoglucose isomerase gene that is subject to carbon source-dependent regulation. J Bacteriol. 1991 May;173(10):3215–3223. doi: 10.1128/jb.173.10.3215-3223.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hsiao C. D., Chou C. C., Hsiao Y. Y., Sun Y. J., Meng M. Expression, purification, and crystallization of two isozymes of 6-phosphoglucose isomerase of Bacillus stearothermophilus. J Struct Biol. 1997 Nov;120(2):196–200. doi: 10.1006/jsbi.1997.3913. [DOI] [PubMed] [Google Scholar]
  10. Huppke P., Wünsch D., Pekrun A., Kind R., Winkler H., Schröter W., Lakomek M. Glucose phosphate isomerase deficiency: biochemical and molecular genetic studies on the enzyme variants of two patients with severe haemolytic anaemia. Eur J Pediatr. 1997 Aug;156(8):605–609. doi: 10.1007/s004310050674. [DOI] [PubMed] [Google Scholar]
  11. Kanno H., Fujii H., Miwa S. Expression and enzymatic characterization of human glucose phosphate isomerase (GPI) variants accounting for GPI deficiency. Blood Cells Mol Dis. 1998 Mar;24(1):54–61. doi: 10.1006/bcmd.1998.0170. [DOI] [PubMed] [Google Scholar]
  12. Lewendon A., Shaw W. V. The pKa of the catalytic histidine residue of chloramphenicol acetyltransferase. Biochem J. 1993 Feb 15;290(Pt 1):15–19. doi: 10.1042/bj2900015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Malaisse W. J., Malaisse-Lagae F., Liemans V., Ottinger R., Willem R. Phosphoglucoisomerase-catalyzed interconversion of hexose phosphates: isotopic discrimination between hydrogen and deuterium. Mol Cell Biochem. 1990 Mar 27;93(2):153–165. doi: 10.1007/BF00226187. [DOI] [PubMed] [Google Scholar]
  14. Meng M., Chen Y. T., Hsiao Y. Y., Itoh Y., Bagdasarian M. Mutational analysis of the conserved cationic residues of Bacillus stearothermophilus 6-phosphoglucose isomerase. Eur J Biochem. 1998 Oct 15;257(2):500–505. doi: 10.1046/j.1432-1327.1998.2570500.x. [DOI] [PubMed] [Google Scholar]
  15. Merkle S., Pretsch W. Glucose-6-phosphate isomerase deficiency associated with nonspherocytic hemolytic anemia in the mouse: an animal model for the human disease. Blood. 1993 Jan 1;81(1):206–213. [PubMed] [Google Scholar]
  16. Niinaka Y., Paku S., Haga A., Watanabe H., Raz A. Expression and secretion of neuroleukin/phosphohexose isomerase/maturation factor as autocrine motility factor by tumor cells. Cancer Res. 1998 Jun 15;58(12):2667–2674. [PubMed] [Google Scholar]
  17. O'Connell E. L., Rose I. A. Affinity labeling of phosphoglucose isomerase by 1,2-anhydrohexitol-6-phosphates. J Biol Chem. 1973 Mar 25;248(6):2225–2231. [PubMed] [Google Scholar]
  18. ROSE I. A., O'CONNELL E. L. Intramolecular hydrogen transfer in the phosphoglucose isomerase reaction. J Biol Chem. 1961 Dec;236:3086–3092. [PubMed] [Google Scholar]
  19. Riordan J. F., McElvany K. D., Borders C. L., Jr Arginyl residues: anion recognition sites in enzymes. Science. 1977 Mar 4;195(4281):884–886. doi: 10.1126/science.190679. [DOI] [PubMed] [Google Scholar]
  20. Rose I. A. Mechanism of the aldose-ketose isomerase reactions. Adv Enzymol Relat Areas Mol Biol. 1975;43:491–517. doi: 10.1002/9780470122884.ch6. [DOI] [PubMed] [Google Scholar]
  21. SALAS M., VINUELA E., SOLS A. SPONTANEOUS AND ENZYMATICALLY CATALYZED ANOMERIZATION OF GLUCOSE 6-PHOSPHATE AND ANOMERIC SPECIFICITY OF RELATED ENZYMES. J Biol Chem. 1965 Feb;240:561–568. [PubMed] [Google Scholar]
  22. Schnackerz K. D., Noltmann E. A. Pyridoxal 5'-phosphate as a site-specific protein reagent for a catalytically critical lysine residue in rabbit muscle phosphoglucose isomerase. Biochemistry. 1971 Dec 21;10(26):4837–4843. doi: 10.1021/bi00802a002. [DOI] [PubMed] [Google Scholar]
  23. Schray K. J., Benkovic S. J., Benkovic P. A., Rose I. A. Catalytic reactions of phosphoglucose isomerase with cyclic forms of glucose 6-phosphate and fructose 6-phosphate. J Biol Chem. 1973 Mar 25;248(6):2219–2224. [PubMed] [Google Scholar]
  24. Sun Y. J., Chou C. C., Chen W. S., Wu R. T., Meng M., Hsiao C. D. The crystal structure of a multifunctional protein: phosphoglucose isomerase/autocrine motility factor/neuroleukin. Proc Natl Acad Sci U S A. 1999 May 11;96(10):5412–5417. doi: 10.1073/pnas.96.10.5412. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Tao W. F., Wang L. L., Shen R. Q., Sheng Z. J. Complete nucleotide sequences of two phosphoglucoisomerase isozymes from Bacillus stearothermophilus. Nucleic Acids Res. 1989 Dec 11;17(23):10107–10108. doi: 10.1093/nar/17.23.10107. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Tekamp-Olson P., Najarian R., Burke R. L. The isolation, characterization and nucleotide sequence of the phosphoglucoisomerase gene of Saccharomyces cerevisiae. Gene. 1988 Dec 15;73(1):153–161. doi: 10.1016/0378-1119(88)90321-6. [DOI] [PubMed] [Google Scholar]
  27. Thomas B. R., Ford V. S., Pichersky E., Gottlieb L. D. Molecular characterization of duplicate cytosolic phosphoglucose isomerase genes in Clarkia and comparison to the single gene in Arabidopsis. Genetics. 1993 Nov;135(3):895–905. doi: 10.1093/genetics/135.3.895. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Watanabe H., Takehana K., Date M., Shinozaki T., Raz A. Tumor cell autocrine motility factor is the neuroleukin/phosphohexose isomerase polypeptide. Cancer Res. 1996 Jul 1;56(13):2960–2963. [PubMed] [Google Scholar]
  29. Xu W., Seiter K., Feldman E., Ahmed T., Chiao J. W. The differentiation and maturation mediator for human myeloid leukemia cells shares homology with neuroleukin or phosphoglucose isomerase. Blood. 1996 Jun 1;87(11):4502–4506. [PubMed] [Google Scholar]

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