Abstract
Prosthetic lipoyl groups are required for the function of several essential multienzyme complexes, such as pyruvate dehydrogenase (PDH), α-ketoglutarate dehydrogenase (KGDH), and the glycine cleavage system (glycine decarboxylase [GDC]). How these proteins are lipoylated has been extensively studied in prokaryotes and yeast (Saccharomyces cerevisiae), but little is known for plants. We earlier reported that mitochondrial fatty acid synthesis by ketoacyl-acyl carrier protein synthase is not vital for protein lipoylation in Arabidopsis (Arabidopsis thaliana) and does not play a significant role in roots. Here, we identify Arabidopsis lipoate-protein ligase (AtLPLA) as an essential mitochondrial enzyme that uses octanoyl-nucleoside monophosphate and possibly other donor substrates for the octanoylation of mitochondrial PDH-E2 and GDC H-protein; it shows no reactivity with bacterial and possibly plant KGDH-E2. The octanoate-activating enzyme is unknown, but we assume that it uses octanoyl moieties provided by mitochondrial β-oxidation. AtLPLA is essential for the octanoylation of PDH-E2, whereas GDC H-protein can optionally also be octanoylated by octanoyltransferase (LIP2) using octanoyl chains provided by mitochondrial ketoacyl-acyl carrier protein synthase to meet the high lipoate requirement of leaf mesophyll mitochondria. Similar to protein lipoylation in yeast, LIP2 likely also transfers octanoyl groups attached to the H-protein to KGDH-E2 but not to PDH-E2, which is exclusively octanoylated by LPLA. We suggest that LPLA and LIP2 together provide a basal protein lipoylation network to plants that is similar to that in other eukaryotes.
Highlights
Prosthetic lipoyl groups are required for the function of several essential multienzyme complexes, such as pyruvate dehydrogenase (PDH), a-ketoglutarate dehydrogenase (KGDH), and the glycine cleavage system
Lipoic acid (LA; 6,8-dithiooctanoic acid) prosthetic groups are essential for the catalytic activity of four important multienzyme complexes in plants and other organisms: pyruvate dehydrogenase (PDH), a-ketoglutarate dehydrogenase (KGDH), branched-chain a-ketoacid dehydrogenase (BCDH), and the Gly cleavage system
We report the identification of the homologous enzyme in Arabidopsis (Arabidopsis thaliana), provide evidence for its mitochondrial location, and show that Arabidopsis lipoate-protein ligase A (LPLA) requires a separate enzyme for octanoate/ lipoate activation
Summary
Prosthetic lipoyl groups are required for the function of several essential multienzyme complexes, such as pyruvate dehydrogenase (PDH), a-ketoglutarate dehydrogenase (KGDH), and the glycine cleavage system (glycine decarboxylase [GDC]). Lipoic acid (LA; 6,8-dithiooctanoic acid) prosthetic groups are essential for the catalytic activity of four important multienzyme complexes in plants and other organisms: pyruvate dehydrogenase (PDH), a-ketoglutarate dehydrogenase (KGDH), branched-chain a-ketoacid dehydrogenase (BCDH), and the Gly cleavage system (glycine decarboxylase [GDC]; Perham, 2000; Douce et al, 2001; Mooney et al, 2002). In all these multienzyme complexes, LA is covalently attached to the «-amino group of a particular lysyl residue of the respective protein subunit. We examine the interplay between LPLA, LIP2, and the mtKAS route of protein lipoylation and suggest a model for protein lipoylation in plant mitochondria
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