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 microtubules, in a concentration-dependent fashion in vitro. The apparent K(d) value of calponin to tubulin was calculated to be 5.2 microM with 2 mol of calponin maximally bound per 1 mol of tubulin. At low ionic strength, tubulin bound to calponin immobilized on Sepharose 4B, and the bound protein was released at about 270 mM NaCl. Chemical cross-linking experiments showed that a 1:1 molar covalent complex of calponin and tubulin was produced. The amount of calponin bound to microtubules decreased with increasing ionic strength or Ca2+ concentration. The addition of calmodulin or S100 to the mixture of calponin and microtubule proteins caused the removal of calponin from microtubules in the presence of Ca2+, but not in the presence of EGTA. Calponin-related proteins 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-treated actin on the binding of calponin to microtubules. Furthermore, only about 20% of calponin-F-actin interaction was inhibited in the presence of an excess amount of tubulin (4 mol per mol of calponin), indicating that tubulin binds to calponin at a different site from that of actin. Compared with MAP2, calponin had little effect on microtubule polymerization.
Fujii
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T
Department of Imagination Science (Kansei Engineering), Faculty of Textile Science and Technology, Shinshu University, Ueda, Nagano.
Hiromori
T
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Hamamoto
M
M
Suzuki
T
T
D000199:Actins / Q000378:metabolism; D017738:Alkanesulfonic Acids; D000818:Animals; D001921:Brain / Q000166:cytology*; D001923:Brain Chemistry; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism; D002148:Calmodulin-Binding Proteins / Q000378:metabolism; D002645:Chickens; D003432:Cross-Linking Reagents; D005022:Ethyldimethylaminopropyl Carbodiimide; D005895:Gizzard, Avian / Q000166:cytology / Q000378:metabolism*; D007700:Kinetics; D008840:Microfilament Proteins; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D008870:Microtubules / Q000378:metabolism*; D009025:Morpholines; D009124:Muscle Proteins / Q000378:metabolism; D009130:Muscle, Smooth / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats; D009418:S100 Proteins / Q000378:metabolism; D012965:Sodium Chloride / Q000494:pharmacology; D014335:Tropomyosin / Q000378:metabolism; D014404:Tubulin / Q000096:biosynthesis / Q000378:metabolism; D000096985:Calponins
D000199:Actins; D017738:Alkanesulfonic Acids; D002135:Calcium-Binding Proteins; D002147:Calmodulin; D002148:Calmodulin-Binding Proteins; D003432:Cross-Linking Reagents; D005022:Ethyldimethylaminopropyl Carbodiimide; D008840:Microfilament Proteins; D008869:Microtubule-Associated Proteins; D009025:Morpholines; D009124:Muscle Proteins; D009418:S100 Proteins; D012965:Sodium Chloride; D014335:Tropomyosin; D014404:Tubulin; D000096985:Calponins; C058992:Tagln protein, mouse; C051615:transgelin; C004550:2-(N-morpholino)ethanesulfonic acid