Identification of palmitoylated mitochondrial proteins using a bio-orthogonal azido-palmitate analogue

CORVI MM; KOSTIUK MA; KELLER BO; PLUMMER G; PRESCHER JA; HANGAUER MJ; BERTOZZI CR; RAJAIAH G; FALCK JR; BERTHIAUME LG

FASEB JOURNAL

FEDERATION AMER SOC EXP BIOL

Año: 2008 vol. 22 p. 721 - 732

0892-6638

Increased levels of circulating saturated free fatty acids such as palmitate have been implicated in the etiology of type II diabetes and cancer. In addition to being a constituent of glycerolipids and a source of energy, palmitate also covalently attaches to numerous cellular proteins via a process named palmitoylation. Recognized for its roles in membrane tethering, cellular signaling and protein trafficking, palmitoylation is also emerging as a potential regulator of metabolism. Indeed, we previously showed that the acylation of two mitochondrial proteins at their active site cysteine residues resulting in their inhibition. Herein, we sought to identify other palmitoylated proteins in mitochondria using a non-radioactive bioorthogonal azido-palmitate analogue that can be selectively derivatized with various tagged-triarylphosphines. Our results show that like palmitate, incorporation of azido-palmitate occurred on mitochondrial proteins via thioester bonds at sites that could be competed by palmitoyl-CoA. Using this method, we identified 21 palmitoylated proteins in the rat liver mitochondrial matrix, a compartment not recognized for its content in palmitoylated proteins and confirmed the palmitoylation of newly identified mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase. We postulate that covalent modification and perhaps inhibition of various mitochondrial enzymes by palmitoyl-CoA could lead to the metabolic impairments found in obesity related diseases.