National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
GM56838
米国
引用
ジャーナル: Biochemistry / 年: 2022 タイトル: Transient Prenyltransferase-Cyclase Association in Fusicoccadiene Synthase, an Assembly-Line Terpene Synthase. 著者: Jacque L Faylo / Trevor van Eeuwen / Kushol Gupta / Kenji Murakami / David W Christianson / 要旨: 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 ...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.
全体 : Octamer of fusicoccadiene synthase with a single interacting cycl...
全体
名称: Octamer of fusicoccadiene synthase with a single interacting cyclase domain, in presence of magnesium cofactor and inhibitor pamidronate
要素
複合体: Octamer of fusicoccadiene synthase with a single interacting cyclase domain, in presence of magnesium cofactor and inhibitor pamidronate
タンパク質・ペプチド: Fusicoccadiene synthase
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超分子 #1: Octamer of fusicoccadiene synthase with a single interacting cycl...
超分子
名称: Octamer of fusicoccadiene synthase with a single interacting cyclase domain, in presence of magnesium cofactor and inhibitor pamidronate タイプ: complex / キメラ: Yes / ID: 1 / 親要素: 0 / 含まれる分子: all