Localization of Cysteine 302 at the Active Site of Aldehyde Dehydrogenase

  • Regina Pietruszko
  • Erich Blatter
  • Darryl P. Abriola
  • Glenn Prestwich
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 284)


The superreactive cysteine was first identified in human cytoplasmic aldehyde dehydrogenase El isozyme, before its primary structure was known, as a part of 35 residue tryptic peptide (Hempel, 1981; Hempel and Pietruszko, 1981; Hempel et al., 1982) by employing iodoacetamide. When the primary structures of the El and E2 isozymes were established (Hempel et al., 1984, 1985; Hsu et al., 1985), this cysteine was found to occupy position 302 in a 500 amino acid residue polypeptide chain. Iodoacetamide fulfilled all criteria for an aldehyde-competitive, active-site-directed reagent with the exception of total inactivation of the mitochondrial E2 isozyme. Since that time, other investigators have also attempted to identify active site residues. Coenzyme-based affinity reagents (von Bahr-Lindstrom et al., 1985) identified cysteines 369 and 302, Nethylmaleimide identified cysteine 49 and 162 (Tu and Weiner, 1988 a,b) and dimethylaminocinnamaldehyde identified serine 74 (Loomes et al., 1990). Our laboratory developed a substrate-based affinity reagent, bromo-acetophenone (MacKerell et al., 1986), which identified glutamate 268 (Abriola et al., 1987).


Tryptic Peptide Aldehyde Dehydrogenase Aspergillus Nidulans Peptide Mapping Radioactive Label 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Abriola, D.P., Fields, R., Stein, S., MacKerell, A.D., Jr. and Pietruszko, R., 1987, Active site of human aldehyde dehydrogenase,Biochemistry, 26:5679.PubMedCrossRefGoogle Scholar
  2. Abriola, D.P., MacKerell, A.D., Jr. and Pietruszko, R., 1990, Correlation of loss of activity of human aldehyde dehydrogenase with reaction of bromoacetophenone with glutamic acid-268 and cysteine-302 residues. Partial-sites reactivity of aldehyde dehydrogenase, Biochem. J., 266:179.PubMedGoogle Scholar
  3. von Bahr-Lindstrom, Hempel, J. and Jornvall, H., 1984, The cytoplasmic isoenzyme of horse liver aldehyde dehydrogenase. Relationship to the corresponding human isoenzyme, Eur. J. Biochem., 141:37.CrossRefGoogle Scholar
  4. von Bahr-Lindstrom, Jeck, R., Woenckhaus, C., Sohn, S., Hempel, J. and Jornvall, H., 1985, Characterization of coenzyme binding site of liver aldehyde dehydrogenase: Differential reactivity of coenzyme analogues, Biochemistry, 24:5847.CrossRefGoogle Scholar
  5. Blatter, E.E., Tasayco, M.L., Prestwich, G. and Pietruszko, R., 1990, Chemical modification of aldehyde dehydrogenase by a vinyl ketone analog of an insect pheromone, Biochem. J., in press.Google Scholar
  6. Carey, F.A. and Sundberg, R.J., 1984, Advanced Organic Chemistry. Second Edition. Part A: Structure and Mechanisms, Plenum Press, New York and London, p. 207.Google Scholar
  7. Ding, Y.-S. and Prestwich, G.D., 1988, Chemical studies of proteins that degrade pheromones: cyclopropanated, fluorinated, and electrophilic analogs of unsaturated aldehyde pheromones, J. Chem. Ecol., 14:2033.CrossRefGoogle Scholar
  8. Dunn, T.J., Koleske, A.J., Lindahl, R. and Pitot, H.C., 1989,Phenobarbital-inducible aldehyde dehydrogenase in the rat. cDNA sequence and regulation of the mRNA by phenobarbital in responsive rats, J. Biol. Chem., 264:13057.PubMedGoogle Scholar
  9. Durst, D., Milano, M., Kikta, E.J., Jr., Connelly, S.A. and Gruska, E., 1975, Phenacyl esters of fatty acids via crown ether catalysts for enhanced ultraviolet detection in liquid chromatography, Analyt. Chem., 47:1797.CrossRefGoogle Scholar
  10. Farres, J., Guan, K.-L. and Weiner, H., 1989, Primary structure of rat and bovine liver mitochondrial aldehyde dehydrogenases deduced from cDNA sequences, Eur. J. Biochem., 180:67.PubMedCrossRefGoogle Scholar
  11. Fukaya, M., Tayama, K., Tamaki, T., Tagami, H., Okumura, H., Kawamura, Y.. and Beppu, T., 1989, Cloning of the membrane-bound aldehyde dehydrogenase gene of Acetobacter polyoxogenesand improvement of acetic acid production by use of the cloned gene, Appl, Environ. Microbiol., 55:171.Google Scholar
  12. Hempel, J.D., 1981, Chemical modification of human liver aldehyde dehydrogenase isoenzymes El and E2. Doctoral Dissertation, Rutgers University. Dissertation Abstracts International 42:3664B, University Microfilms No. DA80204216.Google Scholar
  13. Hempel, J.D. and Pietruszko, R., 1981, Selective chemical modification of human liver aldehyde dehydrogenases El and E2 by iodoacetamide, J. Biol. Chem., 256:10889.PubMedGoogle Scholar
  14. Hempel, J.D., Pietruszko, R., Fietzek, P. andJornvall, H., 1982, Identification of a segment containing a reactive, cysteine residue in human liver cytoplasmic aldehyde dehydrogenase (isoenzyme El)., Biochemistry, 21:6834.PubMedCrossRefGoogle Scholar
  15. Hempel, J., von Bahr-Lindstrom, H. and Jornvall, H., 1984, Aldehyde dehydrogenase from human liver. Primary structure of the cytoplasmic isoenzyme, Eur. J. Biochem., 141:21.PubMedCrossRefGoogle Scholar
  16. Hempel, J., Kaiser, R. and Jornvall, J., 1985, Mitochondrial aldehyde dehydrogenase from human liver. Primary structure, differences in relation to the cytosolic enzyme, and functional correlations, Eur. J. Biochem., 153:13.PubMedCrossRefGoogle Scholar
  17. Hsu, L.C., Tani, K., Fujiyoshi, T., Kurachi, K. and Yoshida, A., 1985, Cloning of cDNAs for human aldehyde dehydrogenases 1 and 2, Proc. Natl. Acad. Sci., USA, 82:3771.PubMedCrossRefGoogle Scholar
  18. Johansson, J., von Bahr-Lindstrom, Jeck, R., Woenckhaus, C. and Jornvall, H., 1988, Mitochondrial aldehyde dehydrogenase from horse liver. Correlations of the same species variants for both the cytosolic and the mitochondrial forms of an enzyme, Eur. J. Biochem., 172:527.PubMedCrossRefGoogle Scholar
  19. Jones, D.E., Jr., Brennan, M.D., Hempel, J. and Lindahl, R., 1988, Cloning and complete nucleotide sequence of a full-length cDNA encoding a catalytically functional tumor-associated aldehyde dehydrogenase, Proc. Natl. Acad. Sci. USA, 85:1782.PubMedCrossRefGoogle Scholar
  20. Kok, M., Oldenhuis, R., van der Linden, M.P.G., Meulenberg, C.H.C., Kingma, J. and Witholt, B., 1989, The Pseudonomas olevoransalkBAC operon encodes two structurally related rubredoxins and an aldehyde dehydrogenase, J. Biol. Chem., 264:5442.PubMedGoogle Scholar
  21. Krzywicki, K.A. and Brandriss, M.C., 1984, Primary structure of the nuclear PUT2 gene involved in the mitochondrial pathway for proline utilization in Saccharomyces cerevisiae, Mol. Cell. Biol., 1984, 4:2837.PubMedGoogle Scholar
  22. Loomes, K.M., Midwinter, G.G.,Blackwell, L.F. and Buckley, P.D., 1990, Evidence for reactivity of serine-74 with trans-4-(N,N-dimethyl- amino)cinnamaldehyde during oxidation by the cytoplasmic aldehyde dehydrogenase from sheep liver, Biochemistry, 29:2070.PubMedCrossRefGoogle Scholar
  23. MacKerell, A.D., Jr., MacWright, R.S. and Pietruszko, R., 1986, Bromoacetophenone as an affinity reagent for human liver aldehyde dehydrogenase, Biochemistry 25:5182.PubMedCrossRefGoogle Scholar
  24. Pedersen, C.J. and Frensdorff, K.H., 1972, Macrocyclic polyethers and their complexes, Angew. Chem., Int. Ed. Engl. 11:16.CrossRefGoogle Scholar
  25. Pickett, M., Gwynne, D.I., Buxton, F.P., Elliott, R., Davies, R.W., Lockington, R.A., Scazzocchio, C. and Sealy-Lewis, H.M., 1987, Cloning and characterization of the aldA gene of Aspergillus idulans, Gene, 51:217.PubMedCrossRefGoogle Scholar
  26. Pietruszko, R. and Yonetani, T., 1980, Aldehyde dehydrogenase from liver, In: Methods in Enzymology, Collowick and Kaplan (Eds.), 71:772.CrossRefGoogle Scholar
  27. Prestwich, G.D., Graham, S. McG., Handley, M., Latli, B., Streinz, L. and Tasayco J., M.L., 1988, Enzymatic processing of pheromones and pheromone analogs, Experientia, 45:267.Google Scholar
  28. Shaw, E., 1970, Chemical modification by active-site-directed reagents. In: The Enzymes (Student Edition), Vol. 1, Structure and Control, Academic Press, p. 91.Google Scholar
  29. Tasayco J., M.L. and Prestwich, G.D., 1990a, A specific affinity reagent to distinguish aldehyde dehydrogenases and oxidases, J. Biochem. Chem., 265:3094.Google Scholar
  30. Tasayco J., M.L. and Prestwich, G.D., 1990b, Aldehyde-oxidizing enzymes in a adult moth: in vitro study of aldehyde metabolism in Heliothis virescens, Arch. Biochem. Biophys., 276:444.CrossRefGoogle Scholar
  31. Tu, G.-C. and Weiner, H., 1988a, Identification of the cysteine residue in the active site of horse liver mitochondrial aldehyde dehydrogenase, J. Biol. Chem., 263:1212.PubMedGoogle Scholar
  32. Tu, G.-C. and Weiner, H., 1988b, Evidence for two distinct active sites on aldehyde dehydrogenase, J. Biol. Chem., 263:1218.PubMedGoogle Scholar
  33. Weretilnyk, E.A. and Hanson, A.D., 1990, Molecular cloning of a plant etaine-aldehyde dehydrogenase, an enzyme implicated in adaptation to alinity and drought, Proc. Natl. Acad. Sci. USA, 87:2745.PubMedCrossRefGoogle Scholar

Copyright information

© Plenum Press, New York 1990

Authors and Affiliations

  • Regina Pietruszko
    • 1
  • Erich Blatter
    • 1
  • Darryl P. Abriola
    • 1
  • Glenn Prestwich
    • 2
  1. 1.Center of Alcohol StudiesRutgers UniversityPiscatawayUSA
  2. 2.Chemistry DepartmentSUNYStony BrookUSA

Personalised recommendations