title: Ca2+-induced conformational changes of spin-labeled g2 chain bound to myosin and the effect of phosphorylation.
abstract: One of the low molecular weight components of myosin, g2, was isolated by alkali treatment of myosin and was chemically modified with a spin label reagent, 4-maleimido-2,2,6,6-tetramethylpiperidinooxyl. The label on g2 showed a rather weakly immobilized ESR spectrum and it was clearly affected by Ca2+; the half-maximal change was at around pCa 4. The spin-labeled g2 was incorporated into myosin by exchange with the intrinsic g2 of myosin in 0.6 M KSCN or 4 M LiC1. The label on g2 became strongly immobilized on association with myosin. Under the conditions used, ESR spectral change due to Ca2+ occurred at two different concentration ranges, which were as low as pCa 8 and at around pCa 4. Phosphorylated g2 was isolated from myosin after the protein kinase [EC 2.1.1.37]-catalyzed phosphorylation of myosin and it was also modified with the maleimide label. Dephosphorylation of the phosphorylated g2 was performed using E. coli alkaline phosphatase [EC 3.1.3.1]. The effects of Ca2+ on the ESR spectra of phosphorylated and dephosphorylated g2 were investigated on the state associated with myosin. A change in the ESR spectrum from strongly immobilized to weakly immobilized states was observed with both g2 chains on the addition of Ca2+. However, the effective concentration  ranges of Ca2+ were quite different; around pCa 4 for the phosphorylated g2 and around pCa 8 for the dephosphorylated g2. The results indicate that g2 undergoes a conformational change at physiological levels of Ca2+ sufficient to saturate troponin, but it does not do so after phosphorylation.
mesh_terms: D000469:Alkaline Phosphatase; D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D004578:Electron Spin Resonance Spectroscopy; D008274:Magnesium / Q000494:pharmacology; D009218:Myosins / Q000378:metabolism*; D009943:Organophosphorus Compounds / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism; D011487:Protein Conformation / Q000187:drug effects; D011494:Protein Kinases; D011817:Rabbits; D013113:Spin Labels
chemical_list: D009943:Organophosphorus Compounds; D010446:Peptide Fragments; D013113:Spin Labels; D011494:Protein Kinases; D000469:Alkaline Phosphatase; D009218:Myosins; D008274:Magnesium; D002118:Calcium