Candida albicans nucleoside-diphosphate kinase: Purification and characterization

Ricardo M. Biondi, Michel Veron, Katherina Walz, Susana Passeron

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Soluble nucleoside-diphosphate kinase (NDP kinase) from Candida albicans was purified to electrophoretic homogeneity and a partial sequence was determined. The enzyme was kinetically and physically characterized. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the purified enzyme revealed a single polypeptide of 17 kDa upon staining, by immunodetection with heterologous and homologous antibodies, and by autoradiography of the phosphorylated enzyme. Furthermore, isoelectric focusing of the purified native enzyme showed a single acidic band (pI 4.5). These results together with a native molecular mass of 98 kDa suggest a hexameric native enzyme composed of identical subunits. Like NDP kinases from other sources, the catalysis involves a phosphoenzyme intermediate that is rapidly formed upon incubation of the enzyme with ATP. The transfer of phosphate from phosphoprotein intermediate to nucleoside diphosphates is equally fast. Kinetic experiments indicated that GTP and ATP had the lowest K(m) compared to UTP, dTTP, and CTP. GDP acted as a preferred acceptor as assessed by kinetic measurements as well as by competition experiments. Experimental data are presented indicating the existence of a membrane-associated NDP kinase. Preliminary characterization of this enzyme suggests that cytosolic and membrane-associated NDP kinases are similar proteins.

Original languageEnglish
Pages (from-to)187-194
Number of pages8
JournalArchives of Biochemistry and Biophysics
Volume323
Issue number1
DOIs
StatePublished - Oct 30 1995
Externally publishedYes

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Nucleoside-Diphosphate Kinase
Candida
Candida albicans
Purification
Enzymes
Phosphotransferases
Adenosine Triphosphate
Heterophile Antibodies
Membranes
Cytidine Triphosphate
Uridine Triphosphate
Kinetics
Diphosphates
Phosphoproteins
Isoelectric Focusing
Molecular mass
Guanosine Triphosphate
Electrophoresis
Autoradiography
Catalysis

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Candida albicans nucleoside-diphosphate kinase : Purification and characterization. / Biondi, Ricardo M.; Veron, Michel; Walz, Katherina; Passeron, Susana.

In: Archives of Biochemistry and Biophysics, Vol. 323, No. 1, 30.10.1995, p. 187-194.

Research output: Contribution to journalArticle

Biondi, Ricardo M. ; Veron, Michel ; Walz, Katherina ; Passeron, Susana. / Candida albicans nucleoside-diphosphate kinase : Purification and characterization. In: Archives of Biochemistry and Biophysics. 1995 ; Vol. 323, No. 1. pp. 187-194.
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