Calreticulin down-regulates both GTP binding and transglutaminase activities of transglutaminase II

Enzyme regulation is an important mechanism for controlling cell proliferat ion and differentiation in response to extracellular signaling molecules. W e have previously reported that a similar to 50 kDa protein (termed G beta( h)) consistently copurified with G alpha(h) (transglutaminase II, TGII) and that G beta(h) down-regulates the GTPase function of TGII by associating w ith GDP-bound TGII [Baek et al. (1996) Biochemistry 35, 2651-2657], In this study, we examined the identity of G beta(h) by partial amino acid sequenc ing and immunological characterizations. The results strongly suggest that G beta(h) is a protein known as calreticulin (CRT). When the regulatory rol e of CRT in the GTPase activity of TGII was examined, CRT inhibited GTP (GT P gamma S) binding and hydrolysis in a concentration-dependent manner. More over, CRT interacted only with GDP-bound TGII. These results demonstrate th at CRT down-regulates the GTPase activity of TGII by associating with GDP-b ound TGII. Studies on the modulation of the TGase activity of TGII revealed that CRT also inhibited TGase activity. The inhibition showed the two char acteristics depend on guanine nucleotides occupying the GTPase active site. The inhibition of the "empty" form of the GTPase active site increased the Ca2+ requirement without changing the V-max. On the other hand, the inhibi tion of the GDP-bound form decreased V-max but did not alter the Ca2+ requi rement. Moreover, the GTP gamma S-bound TGII was virtually resistant to Ca2 +-mediated stimulation of the TGase activity, indicating that the GTP-bound TGII does not function as a TGase. We concluded that CRT is the regulatory protein of TGII that down-regulates both GTPase and TGase activities, oppo sing the activators of TGII function.