Calcium is essential for both the membrane binding and lytic activity of pore-forming protein (perforin) from cytotoxic T-lymphocyte.
The influence of Ca on the membrane binding and lytic activity of lymphocyte pore-forming protein (perforin) was studied. In the absence of Ca, perforin did not bind to the target membranes and did not support lysis of the target cells. In contrast, in the presence of Ca perforin was able to bind to the cell membrane (Km greater than 0.2 mM). Almost all the perforin molecules bind to the membrane within 1 min at 0 degrees C. The addition of EDTA abolished the binding, indicating that the effects of Ca on the membrane binding are reversible. On the other hand, the perforin-mediated lysis of target cells was temp-dependent and also required the presence of Ca in the reaction mixture (Km = 0.05 mM). The difference between the Km values for the membrane binding and lytic activity suggests the presence of two distinct Ca-requiring steps in perforin-mediated target cell lysis.
National Institute of Neuroscience, NCNP, Tokyo, Japan.
S
S
Ishiura
K
K
Matsuda
H
H
Koizumi
T
T
Tsukahara
K
K
Arahata
H
H
Sugita
D000818:Animals; D002118:Calcium / Q000502:physiology*; D002460:Cell Line; D004910:Erythrocyte Membrane / Q000378:metabolism; D006461:Hemolysis; D008562:Membrane Glycoproteins; D008565:Membrane Proteins / Q000502:physiology*; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins; D011485:Protein Binding; D012756:Sheep; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology / Q000502:physiology*; D015032:Zinc / Q000502:physiology
D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D052899:Pore Forming Cytotoxic Proteins; D054353:Perforin; D015032:Zinc; D002118:Calcium