Abstract
Pseudomonas sp. strain AT3 grew with dl-tropic acid, the aromatic component of the alkaloid atropine, as the sole source of carbon and energy. Tropic acid-grown cells rapidly oxidized the growth substrate, phenylacetaldehyde, and phenylacetic acid. Crude cell extracts, prepared from dl-tropic acid-grown cells, contained two NAD+-linked dehydrogenases which were separated by ion-exchange chromatography and shown to be specific for their respective substrates, dl-tropic acid and phenylacetaldehyde. Phenylacetaldehyde dehydrogenase was relatively unstable. The stable tropic acid dehydrogenase was purified to homogeneity by a combination of ion-exchange, molecular-sieve, and affinity chromatography. It had a pH optimum of 9.5 and was equally active with both enantiomers of tropic acid, and at this pH, phenylacetaldehyde was the only detectable product of tropic acid oxidation. The formation of phenylacetaldehyde from tropic acid requires, in addition to dehydrogenation, a decarboxylation step. By analogy with NAD+-specific isocitrate and malate dehydrogenases, phenylmalonic semialdehyde, a 3-oxoacid, would be expected to be the precursor of phenylacetaldehyde. Other workers have established that isocitrate and malate dehydrogenases catalyze the decarboxylation of enzyme-bound or added 3-oxoacid intermediates, a reaction that requires Mn2+ or Mg2+ ions. Studies with tropic acid dehydrogenase were hampered by lack of availability of phenylmalonic semialdehyde, but in the absence of added divalent metal ions, both enantiomers of tropic acid were completely oxidized and we have not, by a number of approaches, found any evidence for the transient accumulation of phenylmalonic semialdehyde.
Full Text
The Full Text of this article is available as a PDF (151.4 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Bartholomew B. A., Smith M. J., Long M. T., Darcy P. J., Trudgill P. W., Hopper D. J. The isolation and identification of 6-hydroxycyclohepta-1,4-dione as a novel intermediate in the bacterial degradation of atropine. Biochem J. 1993 Jul 1;293(Pt 1):115–118. doi: 10.1042/bj2930115. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bartholomew B. A., Smith M. J., Trudgill P. W., Hopper D. J. Atropine Metabolism by Pseudomonas sp. Strain AT3: Evidence for Nortropine as an Intermediate in Tropine Breakdown and Reactions Leading to Succinate. Appl Environ Microbiol. 1996 Sep;62(9):3245–3250. doi: 10.1128/aem.62.9.3245-3250.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
- HATHAWAY J. A., ATKINSON D. E. THE EFFECT OF ADENYLIC ACID ON YEAST NICOTINAMIDE ADENINE DINUCLEOTIDE ISOCITRATE DEHYDROGENASE, A POSSIBLE METABOLIC CONTROL MECHANISM. J Biol Chem. 1963 Aug;238:2875–2881. [PubMed] [Google Scholar]
- KACZKOWSKI J., MOZEJKO-TOCZKO M. [Bacteria decomposing tropane alkaloids]. Acta Microbiol Pol. 1960;9:173–179. [PubMed] [Google Scholar]
- KORKES S., DEL CAMPILLO A., OCHOA S. Biosynthesis of dicarboxylic acids by carbon dioxide fixation. IV. Isolation and properties of an adaptive "malic" enzyme from Lactobacillus arabinosus. J Biol Chem. 1950 Dec;187(2):891–905. [PubMed] [Google Scholar]
- Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
- MOYLE J. Some properties of purified isocitric enzyme. Biochem J. 1956 Aug;63(4):552–558. doi: 10.1042/bj0630552. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Michel H. O., Hackley E., Kramer D. N. Enzymic assay of tropic acid esters. Anal Biochem. 1970 Aug;36(2):294–302. doi: 10.1016/0003-2697(70)90363-5. [DOI] [PubMed] [Google Scholar]
- NIEMER H., BUCHERER H., KOHLER A. [On the decomposition of atropine by Corynebacterium belladonnae]. Hoppe Seylers Z Physiol Chem. 1959;317:238–242. doi: 10.1515/bchm2.1959.317.1.238. [DOI] [PubMed] [Google Scholar]
- NIEMER H., BUCHERER H. [On the degradation of atropine by Corynebacterium belladonnae. II]. Hoppe Seylers Z Physiol Chem. 1961 Oct 25;326:9–12. doi: 10.1515/bchm2.1961.326.1.9. [DOI] [PubMed] [Google Scholar]
- Nascimento A. L., Cilento G. Schiff base formation with amino acids enhances light emission and damage induced in neutrophils by phenylacetaldehyde. Biochim Biophys Acta. 1989 Apr 25;991(1):50–55. doi: 10.1016/0304-4165(89)90027-5. [DOI] [PubMed] [Google Scholar]
- Oosterbaan R. A., Berends F. A comparative study of the atropinesterases of nine Pseudomonas strains. Proc K Ned Akad Wet C. 1971;74(2):158–166. [PubMed] [Google Scholar]
- ROSENBERGER R. F., ELSDEN S. R. The yields of Streptococcus faecalis grown in continuous culture. J Gen Microbiol. 1960 Jun;22:726–739. doi: 10.1099/00221287-22-3-726. [DOI] [PubMed] [Google Scholar]
- Rörsch A., Berends F., Bartlema H. C., Stevens W. F. The isolation and properties of Pseudomonas strains growing on atropine and producing atropinesterase. Proc K Ned Akad Wet C. 1971;74(2):132–147. [PubMed] [Google Scholar]
- Stevens W. F., Rörsch A. The breakdown of tropic acid in Pseudomonas putida strain L. I. Utilization of various substrates; the conversion of tropic acid into phenylacetic acid. Biochim Biophys Acta. 1971 Feb 23;230(2):204–211. doi: 10.1016/0304-4165(71)90205-4. [DOI] [PubMed] [Google Scholar]
- van der Drift A. C., Moes G. W., van der Drift E., Rousseeuw B. A. Comparison of the active sites of atropinesterase and some serine proteases by spin-labeling. Biochemistry. 1985 Sep 24;24(20):5333–5342. doi: 10.1021/bi00341a010. [DOI] [PubMed] [Google Scholar]