Tropomodulin binds two tropomyosins: a novel model for actin filament capping.
Tropomodulin, a tropomyosin-binding protein, caps the slow-growing (pointed) end of the actin filament regulating its dynamics. Tropomodulin, therefore, is important for determining cell morphology, cell movement, and muscle contraction. For the first time we show that one tropomodulin molecule simultaneously binds two tropomyosin molecules in a cooperative manner. On the basis of the tropomodulin solution structure and predicted secondary structure, we introduced a series of point mutations in regions important for tropomyosin binding and actin capping. Capping activity of these mutants was assayed by measuring actin polymerization using pyrene fluorescence. Using direct methods (circular dichroism and native gel electrophoresis) for detecting tropomodulin/tropomyosin binding, we localized the second tropomyosin-binding site to residues 109-144. Despite previous reports that the second binding site is for erythrocyte tropomyosin only, we found that both short nonmuscle and long muscle alpha-tropomyosins bind there as well, though with different affinities. We propose a model for actin capping where one tropomodulin molecule can bind to two tropomyosin molecules at the pointed end.
Kostyukova
Alla S
AS
Department of Neuroscience and Cell Biology, Robert Wood Johnson Medical School, 675 Hoes Lane, Piscataway, New Jersey 08854, USA. kostyuas@umdnj.edu
Choy
Andy
A
Rapp
Brian A
BA
D008841:Actin Cytoskeleton / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D002465:Cell Movement / Q000502:physiology; D004912:Erythrocytes / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D009119:Muscle Contraction / Q000502:physiology; D009132:Muscles / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D051348:Tropomodulin / Q000235:genetics / Q000378:metabolism*; D014335:Tropomyosin / Q000235:genetics / Q000378:metabolism*
D051348:Tropomodulin; D014335:Tropomyosin