Functional Analyses of Two Cellular Binding Domains of Bovine Lactadherin
- 26 April 2000
- journal article
- Published by American Chemical Society (ACS) in Biochemistry
- Vol. 39 (20), 6200-6206
- https://doi.org/10.1021/bi992221r
Abstract
The glycoprotein bovine lactadherin (formerly known as PAS-6/7) comprises two EGF-like domains and two C-like domains found in blood clotting factors V and VIII. Bovine lactadherin binds to αvβ5 integrin in an RGD-dependent manner and also to phospholipids, especially phosphatidyl serine. To define and characterize these bindings the interactions between lactadherin and different mammalian cell types were investigated. Using recombinant forms of bovine lactadherin, the human breast carcinomas MCF-7 cells expressing the αvβ5 integrin receptor were shown to bind specifically to RGD containing lactadherin but not to a mutated RGE lactadherin. A monoclonal antibody against the αvβ5 integrin receptor and a synthetic RGD-containing peptide inhibited the adhesion of MCF-7 cells to lactadherin. Green monkey kidney MA-104 cells, also expressing the αvβ3 together with the αvβ5 integrin, showed binding to bovine lactadherin via both integrins. To investigate the interaction of lipid with lactadherin two fragments were expressed corresponding to the C1C2 domains and the C2 domain. Both fragments bound to phosphatidyl serine in a concentration-dependent manner with an affinity similar to native lactadherin (Kd = 1.8 nM). A peptide corresponding to the C-terminal part of the C2 domain inhibited the binding of lactadherin to phospholipid in a concentration-dependent manner, and finally it was shown that lactadherin mediates binding between artificial phosphatidyl serine membranes and MCF-7 cells. Taken together these results show that lactadherin can act as link between two surfaces by binding to integrin receptors through its N-terminus and to phospholipids through its C-terminus.Keywords
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