Direct structural insight into the substrate-shuttling mechanism of yeast fatty acid synthase by electron cryomicroscopy

Abstract

Yeast fatty acid synthase (FAS) is a 2.6-MDa barrel-shaped multienzyme complex, which carries out cyclic synthesis of fatty acids. By electron cryomicroscopy of single particles we obtained a threedimensional map of yeast FAS at 5.9-Å resolution. Compared to the crystal structures of fungal FAS, the EM map reveals major differences and new features that indicate a considerably different arrangement of the complex in solution compared to the crystal structures, as well as a high degree of variance inside the barrel. Distinct density regions in the reaction chambers next to each of the catalytic domains fitted the substrate-binding acyl carrier protein (ACP) domain. In each case, this resulted in the expected distance of ∼18 Å from the ACP substrate-binding site to the active site of the catalytic domains. The multiple, partially occupied positions of the ACP within the reaction chamber provide direct structural insight into the substrate-shuttling mechanism of fatty acid synthesis in this large cellular machine.cryoelectron microscopy | fatty acid synthesis | acyl carrier protein | molecular architecture

Fatty acid biosynthesis revisited: structure elucidation and metabolic engineering

“…The yeast type I FAS (PDB: 3HMJ) 39 offers a similar barrel shape with a different domain arrangement, further studied by electron microscopy (EM). 40 A larger reaction chamber than the fungal FAS was observed by crystallography, and observed variable domain occupancy by the ACPs further suggests a stochastic role for the ACP within type I complexes. An excellent current review by Grininger discusses type I structure and the evolution of these megasynthases.…”

Structural definition of the lysine swing in Arabidopsis thaliana PDX1: Intermediate channeling facilitating vitamin B 6 biosynthesis

“…Substrate channeling can be mediated by tunneling (22) or by transfer via a covalently bound prosthetic group to the successive active site. Classic examples of the latter are lipoyl-lysine residues of 2-oxo acid dehydrogenases (23), biotinyl-lysine residues of carboxylases (24), and phosphopantetheinyl-serine residues of fatty acid synthases (25,26). Notably, the referred to prosthetic groups are all vitaminderived coenzymes.…”

Fatty acid synthase: insights in the substrate shuttling mechanism by cryo-EM

“…The ACP is tethered by two flexible linkers and shuttles the growing acyl chain between catalytic sites. By electron cryo-microscopy of single particles we obtained a 3D map of yeast FAS at 5.9 Å resolution [1]. Compared to the crystal structures of fungal FAS [2][3][4], the EM map reveals major differences and new features that indicate a considerably different arrangement of the complex in solution compared to the crystal structures.…”

“…Such NTPases have been characterized in detail from both RNA and tailed DNA viruses. We have studied the structure of the non-tailed, membrane-containing, DNA, hyperthermoacidophilic, archaeal virus Sulfolobus turreted icosahedral virus 2 by three-dimensional electron microscopy [1] and now present the crystal structure and activity of its NTPase, B204. The protein binds both single stranded and double stranded nucleic acids, and has an optimum activity at pH 4.5, 80°C.…”

“…The overall fold of B204 places it in the Ftsk-HerA superfamily and sheds light on the evolution of viruses from the PRD1-lineage of viruses. [1] Happonen, L.J. Redder, P., Peng, X., Reigstad, L. Fatty acid synthase (FAS) in yeast and fungi is a 2.6 MDa barrel-shaped multienzyme complex of composition a 6 b 6 with D3 symmetry, which carries out cyclic synthesis of fatty acids from actetyl-and malonyl-CoA.…”