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Structural characterization of recombinant domain II of the basement membrane proteoglycan perlecan

M Costell1, T Sasaki, K Mann

  • 1Max-Planck-Institut für Biochemie, Martinsried, Germany.

FEBS Letters
|November 4, 1996
PubMed

Insights

Researchers purified mouse perlecan domain II, revealing its globular structure and rod-like elements formed by LA modules. This study characterizes the protein

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Structural Biology

Background:

  • Perlecan is a major basement membrane component crucial for tissue development and integrity.
  • Domain II of perlecan, containing LA and IG modules, plays a significant role in perlecan's structure and function.

Purpose of the Study:

  • To characterize the structure and post-translational modifications of mouse perlecan domain II.
  • To investigate the contribution of LA modules to the domain's overall architecture.

Main Methods:

  • Purification of recombinant mouse perlecan domain II from transfected mammalian cells.
  • Rotary shadowing electron microscopy for structural visualization.
  • Circular dichroism (CD) spectroscopy to assess protein folding.
  • Immunological epitope mapping and limited proteolysis to confirm native structure.
  • Analysis of N- and O-glycosylation patterns.

Main Results:

  • Purified perlecan domain II exhibited a globular head connected to a variable-length rod-like segment.
  • Tandem arrays of LA modules were suggested to form the rod-like structures.
  • CD spectroscopy indicated 37% beta structure, and shared epitopes confirmed native folding.
  • A single N-linked glycosylation site and 7-8 O-linked oligosaccharides, primarily in the proline-rich link region, were identified.

Conclusions:

  • Mouse perlecan domain II possesses a distinct structural organization with globular and rod-like features.
  • The LA modules are critical for forming the rod-like elements within the domain.
  • Post-translational modifications, including glycosylation, are important features of perlecan domain II.

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