Cross-linking of lipocortin I and enhancement of its Ca2+ sensitivity by tissue transglutaminase.

The stimulation of human epidermoid carcinoma A431 cells with the calcium ionophore A23187 resulted in the formation of high-molecular-weight lipocortins I, having apparent molecular weights of 75 kDa and 160 kDa as detected with specific anti-lipocortin I antibody. These immunoreactive proteins were identified to be covalently cross-linked multimers of lipocortin I, since essentially the same cross-linked multimers were observed when purified lipocortin I was incubated with tissue transglutaminase (TGase) in vitro. Classical amine substrates for TGase, such as dansylcadaverine and putrescine, were also incorporated stoichiometrically into lipocortin I. Cross-linking or amine incorporation was not observed with lipocortin II. Des 1-26 lipocortin I did not serve as a substrate for TGase, indicating that the N-terminal region of lipocortin I plays an important role in the formation of lipocortin I multimers. The cross-linking of lipocortin I by TGase resulted in a remarkable enhancement of calcium sensitivity for phospholipid binding; i.e., the free calcium concentration required for the cross-linked lipocortin I to attain 50% maximal binding to phosphatidylserine vesicles was as little as 3 microM, while that required for intact monomeric lipocortin I was 20 microM.