Purification and characterization of caldesmon77: a calmodulin-binding protein that interacts with actin filaments from bovine adrenal medulla.
Caldesmon150, a protein composed of the Mr 150,000/147,000 doublet, alternately binds to calmodulin and actin filaments in a Ca2+-dependent "flip-flop" fashion. In all fibroblast cell lines examined, we also found a Mr 77,000 protein that crossreacts with anti-caldesmon150 antibody by using an immunoprecipitation technique [Owada, M.K., Hakura, A., Iida, K., Yahara, I., Sobue, K. & Kakiuchi, S. (1984) Proc. Natl. Acad. Sci. USA 81, 3133-3137]. In this report, we examine the tissue distribution of caldesmon by the method of immunoblotting, using caldesmon-specific antibody. Both caldesmon150 and caldesmon77 show widespread distribution in the tissues examined. Caldesmon77 is more widely distributed than caldesmon150, and we have purified caldesmon77 from bovine adrenal medulla. Its molecular weight estimated by NaDodSO4/polyacrylamide gel electrophoresis was 77,000, and a tetramer of this polypeptide may constitute the native molecule (Mr, 300,000). Caldesmon77 possesses a number of features in common with caldesmon150, including flip-flop binding to calmodulin and actin filaments depending on the concentration of Ca2+ and crossreactivity with caldesmon150-specific antibody. Analysis of caldesmon77-F actin interaction by sedimentation and electrophoresis revealed that 0.5 mg of caldesmon77 bound to 1 mg of F actin. This indicated that the molar ratio between caldesmon77 (tetramer) and actin monomer was calculated to be 1:12-14. In addition, caldesmon77 regulated the actin-myosin interaction in Ca2+-sensitive actomyosin obtained from adrenal medulla. These results suggest that caldesmon77 might be a ubiquitous actin-linked regulator of nonmuscle contractile processes, including those in adrenal medulla.
Sobue
K
K
Tanaka
T
T
Kanda
K
K
Ashino
N
N
Kakiuchi
S
S
D000199:Actins / Q000378:metabolism*; D000313:Adrenal Medulla / Q000378:metabolism; D000818:Animals; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism*; D002148:Calmodulin-Binding Proteins; D002417:Cattle; D003429:Cross Reactions; D007163:Immunosorbent Techniques; D008840:Microfilament Proteins / Q000378:metabolism*; D008970:Molecular Weight; D010749:Phosphoprotein Phosphatases / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D051381:Rats
D000199:Actins; D002135:Calcium-Binding Proteins; D002147:Calmodulin; D002148:Calmodulin-Binding Proteins; D008840:Microfilament Proteins; D010749:Phosphoprotein Phosphatases