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.
Ando
Y
Y
Department of Dermatology, Faculty of Medicine, Kyoto University, Japan.
Imamura
S
S
Owada
M K
MK
Kakunaga
T
T
Kannagi
R
R
D017302:Annexins; D015153:Blotting, Western; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D003432:Cross-Linking Reagents; D004591:Electrophoresis, Polyacrylamide Gel; D004912:Erythrocytes / Q000201:enzymology; D006801:Humans; D011700:Putrescine / Q000378:metabolism; D013379:Substrate Specificity; D011503:Transglutaminases / Q000378:metabolism*; D014407:Tumor Cells, Cultured / Q000187:drug effects
D017302:Annexins; D002135:Calcium-Binding Proteins; D003432:Cross-Linking Reagents; D000001:Calcimycin; D011503:Transglutaminases; D002118:Calcium; D011700:Putrescine