INTERACTION OF CHICKEN GIZZARD SMOOTH-MUSCLE CALPONIN WITH BRAIN MICROTUBULES

Calponin, a major actin-, tropomyosin-, and calmodulin-binding protein in smooth muscle, interacted with tubulin, a main constituent of micr otubules, in a concentration-dependent fashion in, vitro, The apparent K-d value of calponin to tubulin was calculated to be 5.2 mu M with 2 mol of calponin maximally bound per 1 mol of tubulin, At low ionic st rength, tubulin bound to calponin immobilized on Sepharose 4B, and the bound protein was released at about 270 mM NaCl. Chemical cross-linki ng experiments showed that a 1:1 molar covalent complex of calponin an d tubulin was produced, The amount of calponin bound to microtubules d ecreased with increasing ionic strength or Ca2+ concentration. The add ition of calmodulin or S100 to the mixture of calponin and microtubule proteins caused the removal of calponin from microtubules in the pres ence of Ca2+, but not in the presence of EGTA, Calponin-related protei ns including tropomyosin, SM22, and caldesmon had little effect on the calponin binding to microtubules, whereas MAP2 inhibited the binding, Interestingly, there was little, if any, effect of mycalolide B-treat ed actin on the binding of calponin to microtubules, Furthermore, only about 20% of calponin-F-actin interaction was inhibited in the presen ce of an excess amount of tubulin (4 mol per mol of calponin), indicat ing that tubulin binds to calponin at a different site from that of ac tin, Compared with MAP2, calponin had little effect on microtubule pol ymerization.