Intra-mitochondrial poly(ADP-ribosyl)ation: potential role for alpha-ketoglutarate dehydrogenase.
Poly(ADP-ribose) polymerase (PARP) is an intracellular enzyme involved in DNA repair and in building poly-ADP-ribose polymers on nuclear proteins using NAD(+). While the majority of PARP resides in the nucleus, several studies indicated that PARP may also be located in the cytosol or in the mitochondrial matrix. In this study we found several poly-ADP-ribosylated proteins in isolated rat liver mitochondria following hydrogen peroxide (H(2)O(2)) or nitric oxide donor treatment. Protein poly-ADP-ribosylation was more intense in isolated mitochondria than in whole tissue homogenates and it was not associated with increased nuclear PARP activity. We identified five poly-ADP-ribose (PAR) positive mitochondrial bands by protein mass fingerprinting. All of the identified enzymes exhibited decreased activity or decreased levels following oxidative or nitrosative stress. One of the identified proteins is dihydrolipoamide dehydrogenase (DLDH), a component of the alpha-ketoglutarate dehydrogenase (KGDH) complex, which uses NAD(+) as a substrate. This raised the possibility that KGDH may have a PARP-like enzymatic activity. The intrinsic PARP activity of KGDH and DLDH was confirmed using a colorimetric PARP assay kit and by the incubation of the recombinant enzymes with H(2)O(2). The KGDH enzyme may, therefore, have a novel function as a PARP-like enzyme, which may play a role in regulating intramitochondrial NAD(+) and poly(ADP-ribose) homeostasis, with possible roles in physiology and pathophysiology.
Pankotai
Eszter
E
Department of Human Physiology and Clinical Experimental Research, Semmelweis University, Budapest, Hungary.
Lacza
Zsombor
Z
Murányi
Marianna
M
Szabó
Csaba
C
D000818:Animals; D006861:Hydrogen Peroxide / Q000378:metabolism; D007655:Ketoglutarate Dehydrogenase Complex / Q000378:metabolism*; D008099:Liver / Q000201:enzymology / Q000378:metabolism; D008928:Mitochondria / Q000201:enzymology* / Q000378:metabolism*; D008954:Models, Biological; D011064:Poly Adenosine Diphosphate Ribose / Q000378:metabolism*; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D011506:Proteins / Q000378:metabolism*; D051381:Rats
D011506:Proteins; D011064:Poly Adenosine Diphosphate Ribose; D006861:Hydrogen Peroxide; D007655:Ketoglutarate Dehydrogenase Complex; D011065:Poly(ADP-ribose) Polymerases