RRC ID 62697
Author Fujita K, Ohmachi M, Ikezaki K, Yanagida T, Iwaki M.
Title Direct visualization of human myosin II force generation using DNA origami-based thick filaments.
Journal Commun Biol
Abstract The sarcomere, the minimal mechanical unit of muscle, is composed of myosins, which self-assemble into thick filaments that interact with actin-based thin filaments in a highly-structured lattice. This complex imposes a geometric restriction on myosin in force generation. However, how single myosins generate force within the restriction remains elusive and conventional synthetic filaments do not recapitulate the symmetric bipolar filaments in sarcomeres. Here we engineered thick filaments using DNA origami that incorporate human muscle myosin to directly visualize the motion of the heads during force generation in a restricted space. We found that when the head diffuses, it weakly interacts with actin filaments and then strongly binds preferentially to the forward region as a Brownian ratchet. Upon strong binding, the two-step lever-arm swing dominantly halts at the first step and occasionally reverses direction. Our results illustrate the usefulness of our DNA origami-based assay system to dissect the mechanistic details of motor proteins.
Volume 2
Pages 437
Published 2019-1-1
DOI 10.1038/s42003-019-0683-0
PII 683
PMID 31799438
PMC PMC6881340
MeSH Cytoskeletal Proteins / metabolism Cytoskeleton / metabolism Humans Microscopy, Atomic Force Models, Biological Muscle Contraction* Myosin Type II / physiology* Protein Binding Single Molecule Imaging / methods*
DNA material pET T7-7alpha-actinin1-SNAP-His (RDB18384) pShuttle-CMV-human myosin IIa S1 (RDB18385)