Sphingosylphosphocholine reduces the calcium ion requirement for activating tissue transglutaminase

Tissue transglutaminase (tTG) catalyzes a Ca²⁺-dependent transglutaminase reaction resulting in the formation of γ-glutamyl-ε- lysine bonds and is activated during apoptosis to catalyze the formation of apoptotic body. We investigate whether lipids that are membrane components and involved in cell signaling could modify the Ca²⁺-dependent activation of tTG. We found that sphingosylphosphocholine (lyso-SM) was the only lipid to activate transglutaminase at low Ca²⁺ concentrations. In the presence of lyso-SM (125 μM), transglutaminase was detectable at 10 μM Ca²⁺, whereas in the absence of lyso-SM, similar activity was obtained at 160 μM Ca²⁺. Furthermore, in the presence of lipid vesicles lyso-SM retained the ability to enhance the Ca²⁺-dependent activation of tTG. Lyso-SM did not significantly change the K(m) for the glutamyl and primary amine substrates. However, the K(act) for Ca²⁺ was reduced from 300 μM to 90 μM. Structure- function studies of lyso-SM analogs indicate that phosphocholine group on C1, the free amino group at C2 and a C4-C5 double bond are critical for the activation of transglutaminase activity. This is the first demonstration that a specific sphingolipid could enhance the activity of tTG and could play a role in vivo in activation of the tTG at physiologic Ca²⁺ levels.