Crystal structure of the α6β6 holoenzyme of propionyl-coenzyme A carboxylase

Abstract

Propionyl-coenzyme A carboxylase (PCC), a mitochondrial biotin-dependent enzyme, is essential for the catabolism of the amino acids Thr, Val, Ile and Met, cholesterol, and fatty acids with an odd number of carbon atoms. Deficiencies of PCC activity in humans are linked to the disease propionic acidemia (PA), an autosomal recessive disorder that can be fatal in infants 1–4. The holoenzyme of PCC is an α6β6 dodecamer, with a molecular weight of 750 kD. The α subunit contains the biotin carboxylase (BC) and biotin carboxyl carrier protein (BCCP) domains, while the β subunit supplies the carboxyltransferase (CT) activity. Here we report the crystal structure at 3.2 Å resolution of a bacterial PCC α6β6 holoenzyme as well as cryo-electron microscopy (cryo-EM) reconstructionat 15 Å resolution demonstrating a similar structure for human PCC. The structure defines the overall architecture of PCC and reveals unexpectedly that the α subunits are arranged as monomers in the holoenzyme, decorating a central β6 hexamer. A hitherto unrecognized domain in the α subunit, formed by residues between the BC and BCCP domains, is crucial for interactions with the β subunit. We have named it the BT domain. The structure reveals for the first time the relative positions of the BC and CT active sites in the holoenzyme. They are separated by approximately 55 Å, indicating that the entire BCCP domain must translocate during catalysis. The BCCP domain is located in the active site of the β subunit in the current structure, providing insight for its involvement in the CT reaction. The structural information establishes a molecular basis for understanding the large collection of disease-causing mutations in PCC, and also has important relevance for the holoenzymes of other biotin-dependent carboxylases, including 3-methylcrotonyl-CoA carboxylase (MCC) 5–7 and eukaryotic acetyl-CoA carboxylase (ACC) 8,9.

Mechanism for the inhibition of the carboxyltransferase domain of acetyl-coenzyme A carboxylase by pinoxaden

“…2A). This region of the binding site is unlikely to be covered by the BC domain in the full-length ACC enzyme, based on observations on the structure of the propionyl-CoA carboxylase holoenzyme (15).…”

Propionyl Coenzyme A (Propionyl-CoA) Carboxylase in Haloferax mediterranei: Indispensability for Propionyl-CoA Assimilation and Impacts on Global Metabolism

“…PCC is typically composed of three functional components: the biotin carboxylase (BC), the biotin-carboxyl carrier protein (BCCP), and the carboxyltransferase (CT) (6). The crystal structure of a bacterial PCC holoenzyme has been reported, and a similar structure for human PCC has been obtained by using cryo-electron microscopy reconstruction (7). Recently, an acyl-CoA carboxylase with almost equal acetyl-CoA carboxylase (ACC) and PCC activities was characterized for the thermophilic archaeon Metallosphaera sedula (8).…”

Crystal Structure of Urea Carboxylase Provides Insights into the Carboxyltransfer Reaction

“…The BC and BCCP domains of biotin-dependent carboxylases are highly homologous, but their CT domains are distinct. Recently, high resolution crystal structures of the PC (18,19) and PCC (20) holoenzymes have been reported, which greatly advanced the understanding of biotin-dependent carboxylases. However, the molecular mechanism for how UC recognizes the urea substrate and catalyzes its carboxylation is currently not known.…”