Hydrophobic Loop Dynamics and Actin Filament Stability

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• 作者：Damon Scoville ; John D. Stamm ; Dora Toledo-Warshaviak ; Christian Altenbach ; Martin Phillips ; Alexander Shvetsov ; Peter A. Rubenstein ; Wayne L. Hubbell ; Emil Reisler
• 刊名：Biochemistry
• 出版年：2006
• 出版时间：November 14, 2006
• 年：2006
• 卷：45
• 期：45
• 页码：13576 - 13584
• 全文大小：883K
• 年卷期：v.45,no.45(November 14, 2006)
• ISSN：1520-4995

文摘

It has been postulated that the hydrophobic loop of actin (residues 262-274) swings out andinserts into the opposite strand in the filament, stabilizing the filament structure. Here, we analyzed thehydrophobic loop dynamics utilizing four mutants that have cysteine residues introduced at a single locationalong the yeast actin loop. Lateral, copper-catalyzed disulfide cross-linking of the mutant cysteine residuesto the native C374 in the neighboring strand within the filament was fastest for S265C, followed byV266C, L267C, and then L269C. Site-directed spin labeling (SDSL) studies revealed that C265 lies closestto C374 within the filament, followed by C266, C267, and then C269. These results are not predicted bythe Holmes extended loop model of F-actin. Furthermore, we find that disulfide cross-linking destroysL267C and L269C filaments; only small filaments are observed via electron microscopy. Conversely,phalloidin protects the L267C and L269C filaments and inhibits their disulfide cross-linking. Combined,our data indicate that, in solution, the loop resides predominantly in a "parked" position within the filamentbut is able to dynamically populate other conformational states which stabilize or destabilize the filament.Such states may be exploited within a cell by filament-stabilizing and -destabilizing factors.