Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 1994 Jun 1;300(Pt 2):373–381. doi: 10.1042/bj3000373

Investigation of the nature of the two metal-binding sites in 5-aminolaevulinic acid dehydratase from Escherichia coli.

P Spencer 1, P M Jordan 1
PMCID: PMC1138172  PMID: 8002941

Abstract

Two distinct metal-binding sites, termed alpha and beta, have been characterized in 5-aminolaevulinic acid dehydratase from Escherichia coli. The alpha-site binds a Zn2+ ion that is essential for catalytic activity. This site can also utilize other metal ions able to function as a Lewis acid in the reaction mechanism, such as Mg2+ or Co2+. The beta-site is exclusively a transition-metal-ion-binding site thought to be involved in protein conformation, although a metal bound at this site only appears to be essential for activity if Mg2+ is to be bound at the alpha-site. The alpha- and beta-sites may be distinguished from one another by their different abilities to bind divalent-metal ions at different pH values. The occupancy of the beta-site with Zn2+ results in a decrease of protein fluorescence at pH 6. Occupancy of the alpha- and beta-sites with Co2+ results in u.v.-visible spectral changes. Spectroscopic studies with Co2+ have tentatively identified three cysteine residues at the beta-site and one at the alpha-site. Reaction with N-ethyl[14C]maleimide preferentially labels cysteine-130 at the alpha-site when Co2+ occupies the beta-site.

Full text

PDF
377

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anderson P. M., Desnick R. J. Purification and properties of delta-aminolevulinate dehydrase from human erythrocytes. J Biol Chem. 1979 Aug 10;254(15):6924–6930. [PubMed] [Google Scholar]
  2. Bevan D. R., Bodlaender P., Shemin D. Mechanism of porphobilinogen synthase. Requirement of Zn2+ for enzyme activity. J Biol Chem. 1980 Mar 10;255(5):2030–2035. [PubMed] [Google Scholar]
  3. Boese Q. F., Spano A. J., Li J. M., Timko M. P. Aminolevulinic acid dehydratase in pea (Pisum sativum L.). Identification of an unusual metal-binding domain in the plant enzyme. J Biol Chem. 1991 Sep 15;266(26):17060–17066. [PubMed] [Google Scholar]
  4. Christianson D. W. Structural biology of zinc. Adv Protein Chem. 1991;42:281–355. doi: 10.1016/s0065-3233(08)60538-0. [DOI] [PubMed] [Google Scholar]
  5. Dent A. J., Beyersmann D., Block C., Hasnain S. S. Two different zinc sites in bovine 5-aminolevulinate dehydratase distinguished by extended X-ray absorption fine structure. Biochemistry. 1990 Aug 28;29(34):7822–7828. doi: 10.1021/bi00486a007. [DOI] [PubMed] [Google Scholar]
  6. Echelard Y., Dymetryszyn J., Drolet M., Sasarman A. Nucleotide sequence of the hemB gene of Escherichia coli K12. Mol Gen Genet. 1988 Nov;214(3):503–508. doi: 10.1007/BF00330487. [DOI] [PubMed] [Google Scholar]
  7. GIBSON K. D., NEUBERGER A., SCOTT J. J. The purification and properties of delta-aminolaevulic acid dehydrase. Biochem J. 1955 Dec;61(4):618–629. doi: 10.1042/bj0610618. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Gibbs P. N., Chaudhry A. G., Jordan P. M. Purification and properties of 5-aminolaevulinate dehydratase from human erythrocytes. Biochem J. 1985 Aug 15;230(1):25–34. doi: 10.1042/bj2300025. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Gibbs P. N., Jordan P. M. Identification of lysine at the active site of human 5-aminolaevulinate dehydratase. Biochem J. 1986 Jun 1;236(2):447–451. doi: 10.1042/bj2360447. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Glusker J. P. Structural aspects of metal liganding to functional groups in proteins. Adv Protein Chem. 1991;42:1–76. doi: 10.1016/s0065-3233(08)60534-3. [DOI] [PubMed] [Google Scholar]
  11. Jaffe E. K., Abrams W. R., Kaempfen H. X., Harris K. A., Jr 5-Chlorolevulinate modification of porphobilinogen synthase identifies a potential role for the catalytic zinc. Biochemistry. 1992 Feb 25;31(7):2113–2123. doi: 10.1021/bi00122a032. [DOI] [PubMed] [Google Scholar]
  12. Jaffe E. K., Salowe S. P., Chen N. T., DeHaven P. A. Porphobilinogen synthase modification with methylmethanethiosulfonate. A protocol for the investigation of metalloproteins. J Biol Chem. 1984 Apr 25;259(8):5032–5036. [PubMed] [Google Scholar]
  13. Jordan P. M., Gibbs P. N. Mechanism of action of 5-aminolaevulinate dehydratase from human erythrocytes. Biochem J. 1985 May 1;227(3):1015–1020. doi: 10.1042/bj2271015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Jordan P. M., Seehra J. S. 13C NMR as a probe for the study of enzyme-catalysed reactions: mechanism of action of 5-aminolevulinic acid dehydratase. FEBS Lett. 1980 Jun 2;114(2):283–286. doi: 10.1016/0014-5793(80)81134-3. [DOI] [PubMed] [Google Scholar]
  15. Li J. M., Russell C. S., Cosloy S. D. The structure of the Escherichia coli hemB gene. Gene. 1989 Jan 30;75(1):177–184. doi: 10.1016/0378-1119(89)90394-6. [DOI] [PubMed] [Google Scholar]
  16. Liedgens W., Lütz C., Schneider H. A. Molecular properties of 5-aminolevulinic acid dehydratase from Spinacia oleracea. Eur J Biochem. 1983 Sep 1;135(1):75–79. doi: 10.1111/j.1432-1033.1983.tb07619.x. [DOI] [PubMed] [Google Scholar]
  17. Maret W., Andersson I., Dietrich H., Schneider-Bernlöhr H., Einarsson R., Zeppezauer M. Site-specific substituted cobalt(II) horse liver alcohol dehydrogenases. Preparation and characterization in solution, crystalline and immobilized state. Eur J Biochem. 1979 Aug 1;98(2):501–512. doi: 10.1111/j.1432-1033.1979.tb13211.x. [DOI] [PubMed] [Google Scholar]
  18. Mitchell L. W., Jaffe E. K. Porphobilinogen synthase from Escherichia coli is a Zn(II) metalloenzyme stimulated by Mg(II). Arch Biochem Biophys. 1993 Jan;300(1):169–177. doi: 10.1006/abbi.1993.1024. [DOI] [PubMed] [Google Scholar]
  19. Nandi D. L., Baker-Cohen K. F., Shemin D. Delta-aminolevulinic acid dehydratase of Rhodopseudomonas spheroides. J Biol Chem. 1968 Mar 25;243(6):1224–1230. [PubMed] [Google Scholar]
  20. Seehra J. S., Jordan P. M. 5-Aminolevulinic acid dehydratase: alkylation of an essential thiol in the bovine-liver enzyme by active-site-directed reagents. Eur J Biochem. 1981 Jan;113(3):435–446. doi: 10.1111/j.1432-1033.1981.tb05083.x. [DOI] [PubMed] [Google Scholar]
  21. Spencer P., Bown K. J., Scawen M. D., Atkinson T., Gore M. G. Isolation and characterisation of the glycerol dehydrogenase from Bacillus stearothermophilus. Biochim Biophys Acta. 1989 Feb 23;994(3):270–279. doi: 10.1016/0167-4838(89)90304-x. [DOI] [PubMed] [Google Scholar]
  22. Spencer P., Jordan P. M. Purification and characterization of 5-aminolaevulinic acid dehydratase from Escherichia coli and a study of the reactive thiols at the metal-binding domain. Biochem J. 1993 Feb 15;290(Pt 1):279–287. doi: 10.1042/bj2900279. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Spencer P., Scawen M. D., Atkinson T., Gore M. G. The identification of a structurally important cysteine residue in the glycerol dehydrogenase from Bacillus stearothermophilus. Biochim Biophys Acta. 1991 Mar 4;1073(2):386–393. doi: 10.1016/0304-4165(91)90147-9. [DOI] [PubMed] [Google Scholar]
  24. Stinson R. A., Holbrook J. J. Equilibrium binding of nicotinamide nucleotides to lactate dehydrogenases. Biochem J. 1973 Apr;131(4):719–728. doi: 10.1042/bj1310719. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Tsukamoto I., Yoshinaga T., Sano S. The role of zinc with special reference to the essential thiol groups in delta-aminolevulinic acid dehydratase of bovine liver. Biochim Biophys Acta. 1979 Sep 12;570(1):167–178. doi: 10.1016/0005-2744(79)90211-0. [DOI] [PubMed] [Google Scholar]
  26. Wetmur J. G., Bishop D. F., Cantelmo C., Desnick R. J. Human delta-aminolevulinate dehydratase: nucleotide sequence of a full-length cDNA clone. Proc Natl Acad Sci U S A. 1986 Oct;83(20):7703–7707. doi: 10.1073/pnas.83.20.7703. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES