Identification of protein-disulfide isomerase activity in fibronectin

J Biol Chem. 1999 Mar 12;274(11):7032-8. doi: 10.1074/jbc.274.11.7032.

Abstract

Assembly and degradation of fibronectin-containing extracellular matrices are dynamic processes that are up-regulated during wound healing, embryogenesis, and metastasis. Although several of the early steps leading to fibronectin deposition have been identified, the mechanisms leading to the accumulation of fibronectin in disulfide-stabilized multimers are largely unknown. Disulfide-stabilized fibronectin multimers are thought to arise through intra- or intermolecular disulfide exchange. Several proteins involved in disulfide exchange reactions contain the sequence Cys-X-X-Cys in their active sites, including thioredoxin and protein-disulfide isomerase. The twelfth type I module of fibronectin (I12) contains a Cys-X-X-Cys motif, suggesting that fibronectin may have the intrinsic ability to catalyze disulfide bond rearrangement. Using an established protein refolding assay, we demonstrate here that fibronectin has protein-disulfide isomerase activity and that this activity is localized to the carboxyl-terminal type I module I12. I12 was as active on an equal molar basis as intact fibronectin, indicating that most of the protein-disulfide isomerase activity of fibronectin is localized to I12. Moreover, the protein-disulfide isomerase activity of fibronectin appears to be partially cryptic since limited proteolysis of I10-I12 increased its isomerase activity and dramatically enhanced the rate of RNase refolding. This is the first demonstration that fibronectin contains protein-disulfide isomerase activity and suggests that cross-linking of fibronectin in the extracellular matrix may be catalyzed by a disulfide isomerase activity contained within the fibronectin molecule.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Base Sequence
  • Catalysis
  • Chymotrypsin / metabolism
  • DNA Primers
  • Extracellular Matrix / enzymology
  • Extracellular Matrix / metabolism
  • Fibronectins / metabolism*
  • Humans
  • Kinetics
  • Protein Binding
  • Protein Disulfide-Isomerases / metabolism*
  • Protein Folding
  • Ribonucleases / metabolism

Substances

  • DNA Primers
  • Fibronectins
  • Ribonucleases
  • Chymotrypsin
  • Protein Disulfide-Isomerases