UMass Chan Affiliations
Department of Cell BiologyDocument Type
Journal ArticlePublication Date
2005-01-13Keywords
ActinsAllosteric Regulation
Animals
Ca(2+) Mg(2+)-ATPase
Calcium
Cattle
Gelsolin
Macromolecular Substances
Microfilaments
Microscopy, Electron
Muscle, Skeletal
Myosin Subfragments
Particle Size
Protein Binding
Tropomyosin
Troponin
Cell Biology
Metadata
Show full item recordAbstract
Striated muscle thin filaments contain hundreds of actin monomers and scores of troponins and tropomyosins. To study the cooperative mechanism of thin filaments, "mini-thin filaments" were generated by isolating particles nearly matching the minimal structural repeat of thin filaments: a double helix of actin subunits with each strand approximately seven actins long and spanned by a troponin-tropomyosin complex. One end of the particles was capped by a gelsolin (segment 1-3)-TnT fusion protein (substituting for normal TnT), and the other end was capped by tropomodulin. EM showed that the particles were 46 +/- 9 nm long, with a knob-like mass attributable to gelsolin at one end. Average actin, tropomyosin, and gelsolin-troponin composition indicated one troponin-tropomyosin attached to each strand of the two-stranded actin filament. The minifilaments thus nearly represent single regulatory units of thin filaments. The myosin S1 MgATPase rate stimulated by the minifilaments was Ca2+-sensitive, indicating that single regulatory length particles are sufficient for regulation. Ca2+ bound cooperatively to cardiac TnC in conventional thin filaments but noncooperatively to cardiac TnC in minifilaments in the absence of myosin. This suggests that thin filament Ca2+-binding cooperativity reflects indirect troponin-troponin interactions along the long axis of conventional filaments, which do not occur in minifilaments. Despite noncooperative Ca2+ binding to minifilaments in the absence of myosin, Ca2+ cooperatively activated the myosin S1-particle ATPase rate. Two-stranded single regulatory units therefore may be sufficient for myosin-mediated Ca2+-binding cooperativity. Functional mini-thin filaments are well suited for biochemical and structural analysis of thin-filament regulation.Source
Proc Natl Acad Sci U S A. 2005 Jan 18;102(3):656-61. Epub 2005 Jan 11. Link to article on publisher's siteDOI
10.1073/pnas.0407225102Permanent Link to this Item
http://hdl.handle.net/20.500.14038/27665PubMed ID
15644437Related Resources
Link to Article in PubMedae974a485f413a2113503eed53cd6c53
10.1073/pnas.0407225102