Location of heparin-binding sites of fibronectin. Detection of a hitherto unrecognized transamidase sensitive site.
Resolution of a cathepsin D digest of plasma fibronectin on heparin-Sepharose yielded, in addition to non-bound and weakly retained material (Fraction I and II), various fragments which were not eluted until 0.25M NaCl (Fraction III) and 0.5M NaCl (Fraction IV) was applied. Fraction III contained predominantly a peptide of Mr 70 000 originating from the N-terminus of the fibronectin subunits as well as high molecular weight precursors yielding the Mr 70 000 peptide by further digestion. All those peptides were retained by immobilized denatured collagen, type I, indicating the presence of the known gelatin-binding domain. In addition, they contained a transamidase-sensitive site as revealed from a digest of fibronectin previously labelled with [14C]putrescine by a transamidase-mediated reaction. Plasminolysis of the fragment of Mr 70 000 resulted in two peptides of Mr 30 000 and 40 000, only the former being retained by heparin-Sepharose. Fraction IV contained a fragment of Mr 140 000 which, after reduction, dissociated into two peptides of Mr 75 000 and 65 000. Apparently, it included the disulfide bond(s) connecting the two fibronectin subunits close to their C-terminal ends. Partial digestion of the two electrophoretically separated peptide chains with protease of Staphylococcus aureus V8 yielded for each chain a number of peptides with equal electrophoretic migration rate. In addition, however, some peptides were different in the two digests. The results were consistent with an identical or homologous structure of the two peptide chains with an additional sequence in the longer chain. The latter (Mr 75 000) uniquely contained a transamidase susceptible site as demonstrated by processing of [14C]putrescine-labelled fibronectin.
D001665:Binding Sites; D002402:Cathepsin D; D002403:Cathepsins; D005353:Fibronectins / Q000378:metabolism*; D006493:Heparin; D006801:Humans; D008970:Molecular Weight; D010446:Peptide Fragments / Q000032:analysis; D011485:Protein Binding; D000637:Transaminases / Q000378:metabolism*
D005353:Fibronectins; D010446:Peptide Fragments; D006493:Heparin; D000637:Transaminases; D002403:Cathepsins; D002402:Cathepsin D