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Biochem J. Mar 15, 1996; 314(Pt 3): 827–832.
PMCID: PMC1217131

Acidic phospholipids inhibit the intramolecular association between the N- and C-terminal regions of vinculin, exposing actin-binding and protein kinase C phosphorylation sites.

Abstract

Chick vinculin polypeptides expressed in Escherichia coli as glutathione S-transferase (GST) fusion proteins have been used to identify the sites involved in the intramolecular association between the 90 kDa N-terminal head and the 30 kDa C-terminal tail region of the vinculin molecule. Fusion proteins spanning vinculin residues 1-258 and 1-398, immobilized on glutathione-agarose beads, were shown to bind a C-terminal vinculin polypeptide spanning residues 881-1066 (liberated from GST by thrombin cleavage). However, the C-terminal polypeptide did not bind to a fusion protein spanning residues 399-881 or to itself. Binding was dependent on residues 167-207 within the N-terminal polypeptide, a sequence also essential for talin binding. Conversely, the 90 kDa head polypeptide was shown to bind to residues 1029-1036 in the tail region of vinculin. The association of the head and tail was inhibited by acidic, but not neutral, phospholipids. Pre-incubation of vinculin with acidic phospholipids exposed the binding site for F-actin and a phosphorylation site for protein kinase C. The phosphorylation site was located in the tail region of the vinculin molecule. These results raise the possibility that acidic phospholipids play a role in regulating the activity of vinculin and therefore the assembly of both cell-cell and cell-matrix adherens-type junctions.

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Selected References

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