Dynactin 3D Structure: Implications for Assembly and Dynein Binding

JOURNAL OF MOLECULAR BIOLOGY

卷:426

Article

Imai, Hiroshi^1,2  Narita, Akihiro^1,2,3,4,5  Maeda, Yuichiro^1,2,3,4  Schroer, Trina A.⁶ 

[1] Japan Sci & Technol Agcy, RIKEN, ERATo Actin Filament Dynam Project, Sayo, Hyogo 6795148, Japan

[2] RIKEN, Harima Inst SPring Ctr 8, Lab Struct Biochem, Sayo, Hyogo 6795148, Japan

[3] Nagoya Univ, Grad Sch Sci, Struct Biol Res Ctr, Nagoya, Aichi 4648601, Japan

[4] Nagoya Univ, Grad Sch Sci, Div Biol Sci, Nagoya, Aichi 4648601, Japan

[5] JST, PRESTO, Tokyo, Japan

[6] Johns Hopkins Univ, Dept Biol, Baltimore, MD 21218 USA

关键词: dynein;    microtubule;    actin;    single particle analysis;    3D EM;   

DOI  :  10.1016/j.jmb.2014.07.010

来源: Elsevier

【 摘 要 】

The multisubunit protein complex, dynactin, is an essential component of the cytoplasmic dynein motor. High-resolution structural work on dynactin and the dynein/dynactin supercomplex has been limited to small subunits and recombinant fragments that do not report fully on either approximate to 1 MDa assembly. In the present study, we used negative-stain electron microscopy and image analysis based on random conical tilt reconstruction to obtain a three-dimensional (3D) structure of native vertebrate dynactin. The 35-nm-long dynactin molecule has a V-shaped shoulder at one end and a flattened tip at the other end, both offset relative to the long axis of the actin-related protein (Arp) backbone. The shoulder projects dramatically away from the Arp filament core in a way that cannot be appreciated in two-dimensional images, which has implications for the mechanism of dynein binding. The 3D structure allows the helical parameters of the entire Arp filament core, which includes the actin capping protein, CP, to be determined for the first time. This structure exhibits near identity to F-actin and can be well fitted into the dynactin envelope. Molecular fitting of modeled CP-Arp polymers into the envelope shows that the filament contains between 7 and 9 Arp protomers and is capped at both ends. In the 7 Arp model, which agrees best with measured Arp stoichiometry and other structural information, actin capping protein (CP) is not present at the distal tip of the structure, unlike what is seen in the other models. The 3D structure suggests a mechanism for dynactin assembly and length specification. (C) 2014 Elsevier Ltd. All rights reserved.

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