Journal
/
Biochemistry
/
Single-Molecule Analysis of Sf9 Purified Superprocessive Kinesin-3 Family Motors
JoVE Journal
Biochemistry
Author Produced
A subscription to JoVE is required to view this content.  Sign in or start your free trial.
JoVE Journal Biochemistry
Single-Molecule Analysis of Sf9 Purified Superprocessive Kinesin-3 Family Motors

Single-Molecule Analysis of Sf9 Purified Superprocessive Kinesin-3 Family Motors

DOI:
10.3791/63837-v

•

08:16 min

•

July 27, 2022

•

, , ,
1Discipline of Biological Engineering,Indian Institute of Technology Gandhinagar, 2Department of Biotechnology and Bioinformatics,Sambalpur University

Chapters

  • 00:05Introduction
  • 01:22P0 Virus Stock Generation
  • 02:38P1 Virus Stock Generation and Large-Scale Infection
  • 03:28Sf9 Purification of Kinesin-3 Motors
  • 04:25Microtubule Polymerization and In Vitro Single-Molecule Motility Assay
  • 05:22In Vitro Microtubule Gliding Assay
  • 06:23Results: Analysis of Sf9-Purified Kinesin-3 Motor
  • 07:24Conclusion

Summary

Automatic Translation

Automatic Translation

This study details purification of KIF1A(1-393LZ), a member of kinesin-3 family, using Sf9-baculovirus expression system. In vitro single-molecule and multi-motor gliding analysis of these purified motors exhibited robust motility properties comparable to motors from mammalian cell lysate. Thus, Sf9-baculovirus system is amenable to express and purify motor protein of interest.

Tags

Keywords: Sf9 CellsBaculovirus ExpressionSingle-molecule AnalysisKinesin-3 MotorsSuperprocessive MotorsSingle-molecule MotilityMulti-motor Gliding AssaysCytoskeletal ProteinsDionineMyosinProtein PurificationAffinity Purification

Article

Embed

ADD TO PLAYLIST

Usage Statistics

Related Videos

check_url/63837?article_type=v

High Precision FRET at Single-molecule Level for Biomolecule Structure Determination
check_url/63837?article_type=v

Single-molecule Manipulation of G-quadruplexes by Magnetic Tweezers
check_url/63837?article_type=v

Using Three-color Single-molecule FRET to Study the Correlation of Protein Interactions
check_url/63837?article_type=v

Covalent Immobilization of Proteins for the Single Molecule Force Spectroscopy
check_url/63837?article_type=v

Proteome-wide Quantification of Labeling Homogeneity at the Single Molecule Level
check_url/63837?article_type=v

Conventional BODIPY Conjugates for Live-Cell Super-Resolution Microscopy and Single-Molecule Tracking
check_url/63837?article_type=v

An In Vitro Single-Molecule Imaging Assay for the Analysis of Cap-Dependent Translation Kinetics
check_url/63837?article_type=v

Single Cell Analysis Of Transcriptionally Active Alleles By Single Molecule FISH
check_url/63837?article_type=v

Single-Molecule Dwell-Time Analysis of Restriction Endonuclease-Mediated DNA Cleavage
check_url/63837?article_type=v

Single Molecule Fluorescence Energy Transfer Study of Ribosome Protein Synthesis

Read Article