The optical trapping dumbbell assay for nonprocessive motors or motors that turn around filaments

Cold Spring Harb Protoc. 2011 Nov 1;2011(11):1372-4. doi: 10.1101/pdb.prot066688.

Abstract

In vitro motility assays enabled the analysis of coupling between ATP hydrolysis and movement of myosin along actin filaments or kinesin along microtubules. Single-molecule assays using laser trapping have been used to obtain more detailed information about kinesins, myosins, and processive DNA enzymes. The combination of in vitro motility assays with laser-trap measurements has revealed detailed dynamic structural changes associated with the ATPase cycle. This protocol describes the preparation of biotin-actin filaments and coverslips coated with polystyrene beads. These are then used in optical trapping dumbbell assays to study interactions between motors and filaments.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Actin Cytoskeleton / chemistry
  • Actin Cytoskeleton / metabolism*
  • Biotin / chemistry
  • Biotin / metabolism*
  • Microspheres
  • Molecular Motor Proteins / metabolism*
  • Optical Tweezers*
  • Staining and Labeling

Substances

  • Molecular Motor Proteins
  • Biotin