Cloning and kinetic characterization of the Trypanosoma cruzi S-adenosylmethionine decarboxylase

Lisa N. Kinch, Jerry R. Scott, Buddy Ullman, Margaret A. Phillips

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

The gene for S-adenosylmethionine decarboxylase (AdoMetDC), a rate-limiting enzyme in the biosynthesis of polyamines, has been cloned from a Trypanosoma cruz cDNA library. The cDNA clone contains a 1.1 kb open reading frame predicted to encode a 42 kDa protein that shares 31% sequence identity to the human proenzyme. T. cruzi AdoMetDC expressed and purified from E. coli is auto-catalytically processed into two subunits of 32 kDa (α) and 10 kDa (β). The catalytic activity of the purified recombinant enzyme is activated by the addition of putrescine to the reaction. To determine the effect of putrescine on the kinetics of the reaction, the velocity data collected at various substrate and putrescine concentrations were fit to the rate equation describing a non-essential activator. In the presence of fully saturating putrescine, k(cat) increases by 9-fold over the unactivated rate to 0.06 s-1. The model derived K(m) for AdoMet is 0.05 mM in the absence of putrescine and the model-derived K(d) for putrescine binding to free enzyme is 2.5 mM. The K(m) for AdoMet increases by ~2-fold when the enzyme is fully saturated with putrescine. Unlike human AdoMetDC, cadaverine activates the T. cruzi enzyme to a similar extent as putrescine. Copyright (C) 1999 Elsevier Science B.V.

Original languageEnglish (US)
Pages (from-to)1-11
Number of pages11
JournalMolecular and Biochemical Parasitology
Volume101
Issue number1-2
DOIs
StatePublished - Jun 25 1999

Fingerprint

Adenosylmethionine Decarboxylase
Putrescine
Trypanosoma cruzi
Organism Cloning
Enzymes
S-Adenosylmethionine
Cadaverine
Trypanosoma
Enzyme Precursors
Polyamines
Gene Library
Open Reading Frames
Complementary DNA
Clone Cells
Escherichia coli

Keywords

  • Polyamines
  • S-adenosylmethionine decarboxylase
  • T. cruzi

ASJC Scopus subject areas

  • Molecular Biology
  • Parasitology

Cite this

Cloning and kinetic characterization of the Trypanosoma cruzi S-adenosylmethionine decarboxylase. / Kinch, Lisa N.; Scott, Jerry R.; Ullman, Buddy; Phillips, Margaret A.

In: Molecular and Biochemical Parasitology, Vol. 101, No. 1-2, 25.06.1999, p. 1-11.

Research output: Contribution to journalArticle

Kinch, Lisa N. ; Scott, Jerry R. ; Ullman, Buddy ; Phillips, Margaret A. / Cloning and kinetic characterization of the Trypanosoma cruzi S-adenosylmethionine decarboxylase. In: Molecular and Biochemical Parasitology. 1999 ; Vol. 101, No. 1-2. pp. 1-11.
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