D. Seiffert et al., THE SOMATOMEDIN-B DOMAIN OF VITRONECTIN - STRUCTURAL REQUIREMENTS FORTHE BINDING AND STABILIZATION OF ACTIVE TYPE-1 PLASMINOGEN-ACTIVATOR INHIBITOR, The Journal of biological chemistry, 269(4), 1994, pp. 2659-2666
We recently localized the high affinity binding site for activated typ
e plasminogen activator inhibitor (PAI-1) to the somatomedin B domain
(i.e. amino acid (aa) 1-51) of vitronectin (Vn). In this study to furt
her define this site, N-terminal Vn fragments of various lengths were
expressed in Escherichia coli and tested for PAI-1 binding activity. V
n polypeptides containing aa 1-52 and 1-40 retained PAI-1 binding acti
vity and stabilized PAI-1 to a similar extent as intact Vn, but polype
ptides containing aa 1-30 did not bind to PAI-1 nor stabilize its acti
vity. The effects of monoclonal antibodies (mAbs) to Vn on PAI-1 bindi
ng was also determined. One mAb bound to Vn and blocked its ability to
bind to PAI-1. It also dissociated pre-existing PAI-1.Vn complexes, a
nd prevented the incorporation of PAI-1 into extracellular matrix of H
T 1080 cells. This mAb bound to the recombinant peptide containing aa
1-40, but not to the peptide consisting of aa 1-30. A second randomly
chosen mAb with similar affinity for Vn was inactive in these assays a
nd bound to the region between aa 52 and 239. These results indicate t
hat the high affinity binding site for active PAI-1 in Vn is between a
a 1 and 40, and that this domain may also stabilize active PAI-1.