Each actin subunit has three nebulin binding sites: implications for steric blocking.
Nebulin is a giant protein that spans most of the muscle thin filament. Mutations in nebulin result in myopathies and dystrophies. Nebulin contains approximately 200 copies of approximately 35 residue modules, each believed to contain an actin binding site, organized into seven-module superrepeats. The strong correlation between the number of nebulin modules and the length of skeletal muscle thin filaments in different species suggests that nebulin determines thin filament length. Little information exists about the interactions between intact nebulin and F-actin. More insight has come from working with fragments of nebulin, containing from one to hundreds of actin binding modules. However, the observed stoichiometry of binding between these fragments and actin has ranged from 0.4 to 13 modules per actin subunit. We have used electron microscopy and a novel method of helical image analysis to characterize complexes of F-actin with a nebulin fragment. The fragment binds as an extended structure spanning three actin subunits and binding to different sites on each actin. Muscle regulation involves tropomyosin movement on the surface of actin, with binding in three states. Our results suggest the intriguing possibility that intact nebulin may also be able to occupy three different sites on F-actin.
Lukoyanova
Natalya
N
Department of Biochemistry, University of Virginia, Charlottesville 22908-0733, USA.
VanLoock
Margaret S
MS
Orlova
Albina
A
Galkin
Vitold E
VE
Wang
Kuan
K
Egelman
Edward H
EH
D000199:Actins / Q000737:chemistry* / Q000648:ultrastructure; D000494:Allosteric Regulation; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D017124:Conserved Sequence; D006801:Humans; D066298:In Vitro Techniques; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000737:chemistry* / Q000235:genetics / Q000648:ultrastructure; D018482:Muscle, Skeletal / Q000737:chemistry; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000648:ultrastructure; D021122:Protein Subunits; D011817:Rabbits; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000648:ultrastructure; D020449:Repetitive Sequences, Amino Acid
D000199:Actins; D046911:Macromolecular Substances; D009124:Muscle Proteins; D010446:Peptide Fragments; D021122:Protein Subunits; D011994:Recombinant Proteins; C035620:nebulin