Discovery, Structure, and Function of Filamentous 3-Methylcrotonyl-CoA Carboxylase

Abstract

3-methylcrotonyl-CoA carboxylase (MCC) is a biotin-dependent enzyme necessary for leucine catabolism in most organisms. While the crystal structure of recombinant bacterial MCC has been characterized, the structure and potential polymerization of native MCC remain elusive. Here, we discovered that native MCC from Leishmania tarentolae (LtMCC) forms filaments and determined its structure at near-atomic resolution using cryoEM. 6{beta}6 LtMCC dodecamers assemble in a twisted-stacks architecture, manifesting as supramolecular rods extending up to approximately 400 nanometers. LtMCCs in filaments bind biotin but are not covalently biotinylated and lack coenzyme A. Filaments elongate by stacking 6{beta}6 LtMCCs onto the exterior -trimer of the terminal 6{beta}6 dodecamer. This stacking immobilizes the biotin carboxylase domains, sequestering the enzyme in an inactive state within the mitochondrial matrix. Our results support a new model for LtMCC catalysis, termed the dual-swinging-domains model, and cast new light on the functional significance of polymerization in the carboxylase superfamily and beyond.