3 years ago

High-Speed Atomic Force Microscopy Visualization of the Dynamics of the Multienzyme Fatty Acid Synthase

High-Speed Atomic Force Microscopy Visualization of the Dynamics of the Multienzyme Fatty Acid Synthase
Adam Mazur, Habib S. T. Bukhari, Friederike M. C. Benning, Yusuke Sakiyama, Roderick Y. H. Lim, Timm Maier
Multienzymes, such as the protein metazoan fatty acid synthase (FAS), are giant and highly dynamic molecular machines for critical biosynthetic processes. The molecular architecture of FAS was elucidated by static high-resolution crystallographic analysis, while electron microscopy revealed large-scale conformational variability in FAS with some correlation to functional states in catalysis. However, little is known about time scales of conformational dynamics, the trajectory of motions in individual FAS molecules, and the extent of coupling between catalysis and structural changes. Here, we present an experimental single-molecule approach to film immobilized or selectively tethered FAS in solution at different viewing angles and high spatiotemporal resolution using high-speed atomic force microscopy. Mobility of individual regions of the multienzyme is recognized in video sequences, and correlation of shape features implies a convergence of temporal resolution and velocity of FAS dynamics. Conformational variety can be identified and grouped by reference-free 2D class averaging, enabling the tracking of conformational transitions in movies. The approach presented here is suited for comprehensive studies of the dynamics of FAS and other multienzymes in aqueous solution at the single-molecule level.

Publisher URL: http://dx.doi.org/10.1021/acsnano.7b04216

DOI: 10.1021/acsnano.7b04216

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