This Article Full Text Full Text (PDF) All Versions of this Article: biophysj.104.047753v1 88/4/2727    most recent Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Oda, T. Articles by Maéda, Y. Search for Related Content PubMed PubMed Citation Articles by Oda, T. Articles by Maéda, Y.

Position and Orientation of Phalloidin in F-Actin Determined by X-Ray Fiber Diffraction Analysis

Toshiro Oda ^*  , Keiichi Namba  ^¶ and Yuichiro Maéda   ^||

^* Max Planck Institute for Medical Research, Department of Biophysics, Heidelberg, Germany; RIKEN Harima Institute at Spring-8, Laboratory for Structural Biochemistry, Sayo, Hyogo, Japan; Actin Filament Dynamics Project, Exploratory Research for Advanced Technology, Japan Science and Technology Corporation, Sayo, Hyogo, Japan; Dynamic NanoMachine Project, International Cooperative Research Project, Japan Science and Technology Corporation, Suita, Osaka, Japan; ^¶ Graduate School of Frontier Biosciences, Osaka University, Suita, Osaka, Japan; and ^|| Graduate School of Science, Nagoya University, Chikusa, Nagoya, Japan

Correspondence: Address reprint requests to Dr. Toshiro Oda, RIKEN Harima Institute, Kouto 1-1-1, Mikazuki, Sayo 679-5148, Japan. Tel.: 81-791-58-2822; E-mail: toda{at}spring8.or.jp.

Knowledge of the phalloidin binding position in F-actin and the relevant understanding of the mechanism of F-actin stabilization would help to define the structural characteristics of the F-actin filament. To determine the position of bound phalloidin experimentally, x-ray fiber diffraction data were obtained from well-oriented sols of F-actin and the phalloidin-F-actin complex. The differences in the layer-line intensity distributions, which were clearly observed even at low resolution (8 Å), produced well-resolved peaks corresponding to interphalloidin vectors in the cylindrically averaged difference-Patterson map, from which the radial binding position was determined to be 10 Å from the filament axis. Then, the azimuthal and axial positions were determined by single isomorphous replacement phasing and a cross-Patterson map in radial projection to be 84° and 0.5 Å relative to the actin mass center. The refined position was close to the position found by prior researchers. The position of rhodamine attached to phalloidin in the rhodamine-phalloidin-F-actin complex was also determined, in which the conjugated Leu(OH)⁷ residue was found to face the outside of the filament. The position and orientation of the bound phalloidin so determined explain the increase in the interactions between long-pitch strands of F-actin and would also account for the inhibition of phosphate release, which might also contribute to the F-actin stabilization. The method of analysis developed in this study is applicable for the determination of binding positions of other drugs, such as jasplakinolide and dolastatin 11.

This article has been cited by other articles:

				A. Muhlrad, I. Ringel, D. Pavlov, Y. M. Peyser, and E. Reisler Antagonistic Effects of Cofilin, Beryllium Fluoride Complex, and Phalloidin on Subdomain 2 and Nucleotide-Binding Cleft in F-Actin Biophys. J., December 15, 2006; 91(12): 4490 - 4499. [Abstract] [Full Text] [PDF]