Biophysical Society Thematic Meeting | Canterbury 2023

Towards a More Perfect Union: Multi-Scale Models of Muscle and Their Experimental Validation

Monday Speaker Abstracts

STRUCTURES AND MECHANISMS OF ACTIN ATP HYDROLYSIS Yuichiro Maeda 3,6,12 ; Yusuke Kanematsu 1,2 ; Akihiro Narita 3,4 ; Toshiro Oda 5 ; Ryotaro Koike 6 ; Motonori Ota 6 ; Yu Takano 1 ; Kei Moritsugu 7 ; Ikuko Fujiwara 8 ; Kotaro Tanaka 3 ; Hideyuki Komatsu 9 ; Takayuki Nagae 10 ; Nobuhisa Watanabe 10 ; Mitsusada Iwasa 6 ; Shuichi Takeda 3,11 ; 1 Hiroshima City University, Graduate School of Information Sciences, Hiroshima, Japan 2 Hiroshima University, Graduate School of Advanced Science and Engineering, Hiroshima, Japan 3 Nagoya University, Structural Biology Research Center, Nagoya, Japan 4 Nagoya University, Graduate School of Science, Nagoya, Japan 5 Tokai Gakuin University, Faculty of Health and Welfare, Kakamigahara, Japan 6 Nagoya University, Graduate School of Informatics, Nagoya , Japan 7 Yokohama City University, Graduate School of Medical Life Science, Yokohama, Japan 8 Osaka City University, Graduate School of Science, Osaka, Japan 9 Kyushu Institute of Technology, Graduate School of Computer Science and System Engineering, Iizuka, Japan 10 Nagoya University, Synchrotron Radiation Research Center, Nagoya, Japan 11 Okayama University, Research Institute for Interdisciplinary Science, Okayama, Japan 12 Toyota Physical and Chemical Research Institute, Nagakute, Japan The major cytoskeleton protein actin undergoes cyclic transitions between the monomeric G form and the filamentous F-form, which drive organelle transport and cell motility. This mechanical work is driven by the ATPase activity at the catalytic site in the F-form. For deeper understanding of the actin cellular functions, the reaction mechanism must be elucidated. Here, we show that a single actin molecule is trapped in the F-form by fragmin domain-1 binding and present their crystal structures in the ATP analog-, ADP-Pi-, and ADP-bound forms, at 1.15-Å resolutions. By applying quantum mechanics/molecular mechanics calculations to the structures, we have revealed a consistent and comprehensive reaction path of ATP hydrolysis by the F-form actin. The reaction path consists of four steps: 1) W1 and W2 rotations; 2) P G –O 3B bond cleavage; 3) four concomitant events: W1–PO 3 - formation, OH - and proton cleavage, nucleophilic attack by the OH - against P G , and the abstracted proton transfer; and 4) proton relocation that stabilizes the ADP-Pi–bound F-form actin. The mechanism explains how the G to-F conformational transition triggers ATP hydrolysis, the slow rate of ATP hydrolysis by actin, and the irreversibility of the hydrolysis reaction. Remarkably, while the actin active site structure is much simpler than those of some motor proteins, like myosin, actin and the motor proteins share the common reaction path. This common mechanism is now robust, since with actin both the initial and the final structures of the ATP hydrolysis reaction were available at high resolutions for the first time among ATPase proteins. This is made possible due to a unique property of actin; with actin, the end of ATP hydrolysis is not associated with a global protein conformational change. This makes the ADP-Pi-bound state stable. With actin, Pi-release could be due to structural fluctuations, not due to a global conformational change.

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