Functional effects of protein kinases and peroxynitrite on cardiac carnitine palmitoyltransferase-1 in isolated mitochondria

Abstract

We have previously shown that metoprolol can inhibit carnitine palmitoyltransferase-1 catalytic activity and decrease its malonyl CoA sensitivity within 30 min, suggesting the importance of a covalent modification. The aim of this study was to characterize the effects of PTMs on CPT-1 in the heart. Mitochondria were isolated from the hearts of male Wistar rats and incubated with kinases of interest (protein kinase A, CAMK-II, p38 MAPK, Akt) or with peroxynitrite and sodium nitroprusside. PKA decreased CPT-1 malonyl CoA sensitivity, associated with phosphorylation of CPT-1A, whereas CAMK-II increased malonyl CoA sensitivity by phosphorylating CPT-1B. p38 bound to CPT-1B and stimulated CPT-1 activity. The association of CPT-1 with these kinases and their scaffolding proteins was confirmed in co-localization studies. Peroxynitrite and sodium nitroprusside reversibly stimulated CPT-1 activity, and the change in CPT-1B activity was most consistently associated with glutathiolation of CPT-1B. These studies have identified a new regulatory system of kinases, scaffolding proteins and thiol redox chemistry which can control cardiac CPT-1 in vitro.