Domain IV of Mouse Laminin beta1 and beta2 Chains

Eur J Biochem. 2002 Jan;269(2):431-42. doi: 10.1046/j.0014-2956.2001.02663.x.

Abstract

Domain IV, consisting of about 230 residues, represents a particular protein module so far found only in laminin beta1 and beta2 chains. Both domains were obtained by recombinant production in mammalian cells. They showed a globular structure, as expected from electron microscopic examination of laminins. Fragment beta1IV was obtained as a monomer and a disulfide-bonded dimer, and both were modified to approximately 50% by a single chondroitin sulfate chain attached to Ser721 of an SGD consensus sequence. Dimerization is caused by an odd number of cysteines, with three of them having a partial thiol character. Whether both modifications also occur in tissue forms of laminin remains to be established. Fragment beta2IV was only obtained as a monomer, as it lacked one crucial cysteine and the SGD sequence. It required, however, the presence of two adjacent LE modules for proper folding. Polyclonal antibodies raised against both fragments showed no cross-reaction with each other and allowed establishment of beta chain-specific radioimmunoassays and light and electron microscopic immunostaining of tissues. This demonstrated a 5-25-fold lower content of beta2 compared with beta1 chains in various tissue extracts of adult mice. Tissues derived from beta2-deficient mice failed to react with the beta2-specific antibodies but showed a twofold higher content of beta1 than heterozygotes. The antibodies to beta2 showed broader tissue staining than reported previously, including in particular a distinct reaction with the extrasynaptic endomysium of skeletal muscle. Immunogold staining localized both beta chains primarily to basement membranes of kidney, muscle and various other tissues.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • DNA Primers
  • Glycosaminoglycans / chemistry*
  • Immunohistochemistry
  • Laminin / chemistry*
  • Laminin / isolation & purification
  • Laminin / metabolism
  • Mice
  • Molecular Sequence Data
  • Protein Conformation
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / metabolism
  • Sequence Homology, Amino Acid

Substances

  • DNA Primers
  • Glycosaminoglycans
  • Laminin
  • Recombinant Proteins