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Isolation of Intermediate Filament Proteins from Multiple Mouse Tissues to Study Aging-associated Post-translational Modifications
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Isolation of intermediate filaments

Conrad L Leung1, Ronald K H Liem

  • 1Columbia University, New York, New York, USA.

Current Protocols in Cell Biology
|January 30, 2008
PubMed

Insights

This study details methods for purifying intermediate filament (IF) proteins, including neurofilament proteins (NFL, NFM, NFH), from biological sources and recombinant systems for cell biology research.

Area of Science:

  • Cell Biology
  • Biochemistry
  • Structural Biology

Background:

  • Intermediate filaments (IFs) are crucial cytoskeletal components found in eukaryotic cells, composed of diverse IF proteins.
  • IF proteins are typically insoluble, requiring denaturing conditions for purification, followed by reassembly via dialysis.

Purpose of the Study:

  • To provide detailed protocols for isolating and purifying intermediate filament proteins.
  • To illustrate IF purification using neurofilament proteins (NFL, NFM, NFH) from bovine spinal cord.
  • To present an alternative method for purifying NFL from a prokaryotic expression system.

Main Methods:

  • Purification of neurofilament proteins (NFL, NFM, NFH) from bovine spinal cord.
  • Reassembly of purified IF proteins into filaments through dialysis.
  • Purification of NFL from a prokaryotic expression system.

Main Results:

  • Established protocols for isolating native and recombinant intermediate filament proteins.
  • Demonstrated the feasibility of reassembling purified IF proteins into functional filaments.
  • Highlighted the efficiency of prokaryotic expression systems for producing IF proteins.

Conclusions:

  • Detailed purification protocols are essential for studying intermediate filament protein structure and function.
  • Recombinant protein expression offers a straightforward and potentially preferred method for IF purification.
  • Understanding IF purification is key to advancing research in cell structure and dynamics.

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