Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is pyruvylated during 3-bromopyruvate mediated cancer cell death.
BACKGROUND
The pyruvic acid analog 3-bromopyruvate (3BrPA) is an alkylating agent known to induce cancer cell death by blocking glycolysis. The anti-glycolytic effect of 3BrPA is considered to be the inactivation of glycolytic enzymes. Yet, there is a lack of experimental documentation on the direct interaction of 3BrPA with any of the suggested targets during its anticancer effect.


RESULTS
In the current study, using radiolabeled ((14)C) 3BrPA in multiple cancer cell lines, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was identified as the primary intracellular target of 3BrPA, based on two-dimensional (2D) gel electrophoretic autoradiography, mass spectrometry and immunoprecipitation. Furthermore, in vitro enzyme kinetic studies established that 3BrPA has marked affinity to GAPDH. Finally, Annexin V staining and active caspase-3 immunoblotting demonstrated that apoptosis was induced by 3BrPA.


CONCLUSIONS
GAPDH pyruvylation by 3BrPA affects its enzymatic function and is the primary intracellular target in 3BrPA mediated cancer cell death.
Ganapathy-Kanniappan
Shanmugasundaram
S
Russell H. Morgan Department of Radiology and Radiological Sciences, Baltimore, MD 21287, USA.
Geschwind
Jean-Francois H
JF
Kunjithapatham
Rani
R
Buijs
Manon
M
Vossen
Josephina A
JA
Tchernyshyov
Irina
I
Cole
Robert N
RN
Syed
Labiq H
LH
Rao
Pramod P
PP
Ota
Shinichi
S
Vali
Mustafa
M
D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy / Q000201:enzymology / Q000473:pathology*; D015180:Electrophoresis, Gel, Two-Dimensional; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D008113:Liver Neoplasms / Q000188:drug therapy / Q000201:enzymology / Q000473:pathology*; D011773:Pyruvates / Q000494:pharmacology*; D011817:Rabbits; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014407:Tumor Cells, Cultured
D011773:Pyruvates; C017092:bromopyruvate; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases