Identification, Purification, and Characterization of an Eukaryotic-like Phosphopantetheine Adenylyltransferase (Coenzyme A Biosynthetic Pathway) in the Hyperthermophilic Archaeon Pyrococcus abyssi - Archive ouverte HAL Accéder directement au contenu
Article Dans Une Revue Journal of Biological Chemistry Année : 2003

Identification, Purification, and Characterization of an Eukaryotic-like Phosphopantetheine Adenylyltransferase (Coenzyme A Biosynthetic Pathway) in the Hyperthermophilic Archaeon Pyrococcus abyssi

Résumé

Although coenzymeA (CoA) is essential in numerous metabolic pathways in all living cells, molecular characterization of the CoA biosynthetic pathway in Archaea remains undocumented. Archaeal genomes contain detectable homologues for only three of the five steps of the CoA biosynthetic pathway characterized in Eukarya and Bacteria. In case of phosphopantetheine adenylyltransferase (PPAT) (EC 2.7.7.3), the putative archaeal enzyme exhibits significant sequence similarity only with its eukaryotic homologs, an unusual situation for a protein involved in a central metabolic pathway. We have overexpressed in Escherichia coli, purified, and characterized this putative PPAT from the hyperthermophilic archaeon Pyrococcus abyssi (PAB0944). Matrixassisted laser desorption ionization-time of flight mass spectrometry and high performance liquid chromatography measurements are consistent with the presence of a dephospho-CoA (dPCoA) molecule tightly bound to the polypeptide. The protein indeed catalyzes the synthesis of dPCoA from 4-phosphopantetheine and ATP, as well as the reverse reaction. The presence of dPCoA stabilizes PAB0944, as it induces a shift from 76 to 82°C of the apparent T m measured by differential scanning microcalorimetry. Potassium glutamate was found to stabilize the protein at 400 mM. The enzyme behaves as a monomeric protein. Although only distantly related, secondary structure prediction indicates that archaeal and eukaryal PPAT belong to the same nucleotidyltransferase superfamily of bacterial PPAT. The existence of operational proteins highly conserved between Archaea and Eukarya involved in a central metabolic pathway challenge evolutionary scenarios in which eukaryal operational proteins are strictly of bacterial origin.
Fichier principal
Vignette du fichier
Armengaud2003_JBC.pdf (514 Ko) Télécharger le fichier
Origine : Fichiers éditeurs autorisés sur une archive ouverte

Dates et versions

hal-03551632 , version 1 (01-02-2022)

Identifiants

Citer

Jean Armengaud, Bernard Fernandez, Valérie Chaumont, Françoise Rollin-Genetet, Stéphanie Finet, et al.. Identification, Purification, and Characterization of an Eukaryotic-like Phosphopantetheine Adenylyltransferase (Coenzyme A Biosynthetic Pathway) in the Hyperthermophilic Archaeon Pyrococcus abyssi. Journal of Biological Chemistry, 2003, 278 (33), pp.31078 - 31087. ⟨10.1074/jbc.m301891200⟩. ⟨hal-03551632⟩
28 Consultations
44 Téléchargements

Altmetric

Partager

Gmail Facebook X LinkedIn More