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Molecular and Cellular Biochemistry

, Volume 195, Issue 1–2, pp 55–64 | Cite as

Characterization of ornithine decarboxylase and regulation by its antizyme in Thermus thermophilus

  • A.A. Pantazaki
  • C.G. Anagnostopoulos
  • E.E. Lioliou
  • D.A. Kyriakidis
Article

Abstract

Ornithine decarboxylase (ODC), the key enzyme of polyamine biosynthesis was highly purified from the thermophilic bacterium Thermus thermophilus. The enzyme preparation showed a single band on SDS-polyacrylamide gel electrophoresis, a pH optimum of 7.5 and a temperature optimum at 60°C. The native enzyme which is phosphorylated could, upon treatment with alkaline phosphatase, lose all activity. The inactive form could be reversibly activated by nucleotides in the order of NTP>NDP>NMP. When physiological polyamines were added to the purified enzyme in vitro, spermine or spermidine activated ODC by 140 or 40%, respectively, while putrescine caused a small inhibition. The basic amino acids lysine and arginine were competitive inhibitors of ODC, while histidine did not affect the enzyme activity. Among the phosphoamino acids tested, phosphoserine was the most effective activator of purified ODC. Polyamines added at high concentration to the medium resulted in a delay or in a complete inhibition of the growth of T. thermophilus, and in a decrease of the specific activity of ornithine decarboxylase. The decrease of ODC activity resulted from the appearance of a non-competitive inhibitor of ODC, the antizyme (Az). The T. thermophilus antizyme was purified by an ODC-Sepharose affinity column chromatography, as well as by immunoprecipitation using antibodies raised against the E. coli antizyme. The antizyme of E. coli inhibited the ODC of T. thermophilus, and vice versa. The fragment of amino acids 56-292 of the E. coli antizyme, produced as a fusion protein of glutathione S-transferase, did not inhibit the ODC of E. coli or T. thermophilus.

ornithine decarboxylase antizyme Thermus thermophilus 

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Copyright information

© Kluwer Academic Publishers 1999

Authors and Affiliations

  • A.A. Pantazaki
    • 1
  • C.G. Anagnostopoulos
    • 1
  • E.E. Lioliou
    • 1
  • D.A. Kyriakidis
    • 1
  1. 1.Laboratory of Biochemistry, Department of ChemistryAristotle University of ThessalonikiThessalonikiGreece

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