Intracellular Targets of Paullones. Identification following affinity purification on immobilized inhibitor.
Numerous inhibitors of cyclin-dependent kinases and glycogen synthase kinase-3 (GSK-3) are being developed in view of their potential applications against cancers and neurodegenerative disorders. Among these, paullones constitute a family of potent and apparently selective cyclin-dependent kinase and GSK-3 inhibitors. However, their actual intracellular targets remain to be identified. To address this issue we have immobilized a paullone, gwennpaullone, on an agarose matrix. Extracts from various cell types and tissues were screened for proteins interacting with this matrix. This approach validated GSK-3alpha and GSK-3beta as major intracellular paullone targets and also mitochondrial, but not cytoplasmic, malate dehydrogenase (MDH). Mitochondrial MDH was indeed inhibited by micromolar concentrations of paullones. Mitochondrial MDH was the major paullone-binding protein in the parasitic protozoon Leishmania mexicana, and paullones inhibited growth of the parasite. This simple batchwise affinity chromatography approach constitutes a straightforward method for the identification of intracellular targets of this particular class of novel anti-mitotic compounds. It has revealed an unexpected target, mitochondrial MDH, the inhibition of which may participate in the pharmacological effects of paullones.
Knockaert
Marie
M
Station Biologique de Roscoff, CNRS, BP 74, Bretagne, France. knockaer@sb-roscoff.fr
Wieking
Karen
K
Schmitt
Sophie
S
Leost
Maryse
M
Grant
Karen M
KM
Mottram
Jeremy C
JC
Kunick
Conrad
C
Meijer
Laurent
L
D000595:Amino Acid Sequence; D000818:Animals; D001381:Azepines / Q000302:isolation & purification / Q000494:pharmacology*; D001552:Benzazepines; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors*; D002846:Chromatography, Affinity; D004791:Enzyme Inhibitors / Q000302:isolation & purification / Q000494:pharmacology*; D038362:Glycogen Synthase Kinase 3; D038341:Glycogen Synthase Kinases; D007211:Indoles / Q000302:isolation & purification / Q000494:pharmacology*; D007894:Leishmania mexicana / Q000201:enzymology; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid
D001381:Azepines; D001552:Benzazepines; D004791:Enzyme Inhibitors; D007211:Indoles; C462503:gwennpaullone; D038341:Glycogen Synthase Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D038362:Glycogen Synthase Kinase 3