8EAX

Octameric prenyltransferase domain of fusicoccadiene Synthase with C2 symmetry sans transiently associating cyclase domains

Summary for 8EAX

• Entry DOI: 10.2210/pdb8eax/pdb
• EMDB information: 27989
• Descriptor: Fusicoccadiene synthase (1 entity in total)
• Functional Keywords: terpene synthase, prenyltransferase, transferase
• Biological source: Diaporthe amygdali
• Total number of polymer chains: 8
• Total formula weight: 671423.82

Authors

Faylo, J.L.,van Eeuwen, T.,Christianson, D.W. (deposition date: 2022-08-29, release date: 2022-11-02, Last modification date: 2024-06-19)

Primary citation

Faylo, J.L.,van Eeuwen, T.,Gupta, K.,Murakami, K.,Christianson, D.W.
Transient Prenyltransferase-Cyclase Association in Fusicoccadiene Synthase, an Assembly-Line Terpene Synthase.
Biochemistry, 61:2417-2430, 2022

PubMed Abstract: Fusicoccadiene synthase from the fungus (PaFS) is an assembly-line terpene synthase that catalyzes the first two steps in the biosynthesis of Fusiccocin A, a diterpene glycoside. The C-terminal prenyltransferase domain of PaFS catalyzes the condensation of one molecule of C dimethylallyl diphosphate and three molecules of C isopentenyl diphosphate to form C geranylgeranyl diphosphate, which then transits to the cyclase domain for cyclization to form fusicoccadiene. Previous structural studies of PaFS using electron microscopy (EM) revealed a central octameric prenyltransferase core with eight cyclase domains tethered in random distal positions through flexible 70-residue linkers. However, proximal prenyltransferase-cyclase configurations could be captured by covalent cross-linking and observed by cryo-EM and mass spectrometry. Here, we use cryo-EM to show that proximally configured prenyltransferase-cyclase complexes are observable even in the absence of covalent cross-linking; moreover, such complexes can involve multiple cyclase domains. A conserved basic patch on the prenyltransferase domain comprises the primary touchpoint with the cyclase domain. These results support a model for transient prenyltransferase-cyclase association in which the cyclase domains of PaFS are in facile equilibrium between proximal associated and random distal positions relative to the central prenyltransferase octamer. The results of biophysical measurements using small-angle X-ray scattering, analytical ultracentrifugation, dynamic light scattering, and size-exclusion chromatography in-line with multi-angle light scattering are consistent with this model. This model accordingly provides a framework for understanding substrate transit between the prenyltransferase and cyclase domains as well as the cooperativity observed for geranylgeranyl diphosphate cyclization.

PubMed: 36227241
DOI: 10.1021/acs.biochem.2c00509

Experimental method

ELECTRON MICROSCOPY (3.73 Å)