REGULATION OF BRAIN M-CALPAIN CA2-LIPIDS - ACTIVATION AT INTRACELLULAR CA2+ LEVEL( SENSITIVITY BY MIXTURES OF MEMBRANE)

Combinations of certain phospholipids and gangliosides increase the sp ecific activity of m calpain and can activate m calpain at 1 to 10 mu M Ca2+ concentration. However, this level of calcium is still greater than the normal intracellular calcium level, We have used combinations of lipids to demonstrate the m calpain activity at the physiological Ca2+ level, GD1a (100 mu M) and cerebroside (Cerb; 750 mu M; 1:7.5) mi xture was the most effective, At 0.5 mu M to 1.0 mu M Ca2+ concentrati ons, 15-20% of the maximal activity was detected for the purified myel in and cytosolic m calpains, Other combinations were GD1a (100 mu M), GM1 (100 mu M), Cerb (750 mu M), sulfatide (Sulf; 750 mu M), and phosp hatidylinositol (PI; 300 mu M) at a ratio of 1:1:7.5:7.5:3, respective ly, These lipid mixtures stimulated calpain activity at three- to tenf old less calcium concentration than control, The other mixtures, inclu ding GD1a:Sulf (1:9) > GD1a:PI (1:4) > PI:Sulf (1:5) > Cerb:Sulf (1:5) and PI:Cerb (1:2.5), also stimulated calpain activity at 1.0 mu M Ca2 + concentration. Triton X-100, oxidized glutathione (GSSG), and calpai n activator did not affect the Ca2+ requirement. Liposomes containing GD1a, Cerb, and m calpain also showed recognizable calpain activity at a significantly reduced Ca2+ concentration (0.4 mu M), confirming the glycolipid-mediated enzyme modulation, These studies indicate that sp ecific lipid mixtures can stimulate m calpain activity at an intracell ular level of Ca2+. (C) 1996 Wiley-Liss, Inc.