Binding of myotrophin/V-1 to actin-capping protein: Implications for how capping protein binds to the filament barbed end

Nandini Bhattacharya, Shatadal Ghosh, David Sept, John A. Cooper

Research output: Contribution to journalArticle

32 Scopus citations

Abstract

The heterodimeric actin-capping protein (CP) regulates actin assembly and cell motility by binding tightly to the barbed end of the actin filament. Here we demonstrate that myotrophin/V-1 binds directly to CP in a 1:1 molar ratio with a Kd of 10-50 nM. V-1 binding inhibited the ability of CP to cap the barbed ends of actin filaments. The actin-binding COOH-terminal region, the "tentacle," of the CP β subunit was important for binding V-1, with lesser contributions from the α subunit COOH-terminal region and the body of the protein. V-1 appears to be unable to bind to CP that is on the barbed end, based on the observations that V-1 had no activity in an uncapping assay and that the V-1·CP complex had no capping activity. Two loops of V-1, which extend out from the α-helical backbone of this ankyrin repeat protein, were necessary for V-1 to bind CP. Parallel computational studies determined a bound conformation of the β tentacle with V-1 that is consistent with these findings, and they offered insight into experimentally observed differences between the α1 and α2 isoforms as well as the mutant lacking the α tentacle. These results support and extend our "wobble" model for CP binding to the actin filament, in which the two COOH-terminal regions of CP bind independently to the actin filament, and bound CP is able to wobble when attached only via its mobile β-subunit tentacle. This model is also supported by molecular dynamics simulations of CP reported here. The existence of the wobble state may be important for actin dynamics in cells.

Original languageEnglish
Pages (from-to)31021-31030
Number of pages10
JournalJournal of Biological Chemistry
Volume281
Issue number41
DOIs
StatePublished - Oct 13 2006

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