The preparation of calmodulins from barley (Hordeum sp.) and basidiomycete fungi

Abstract

Calmodulin-like proteins were purified from the fruiting bodies of higher (basidiomycete) fungi [Russula spp., Cortinarius spp. and Agaricus bisporus] and barley (Hordeum sp.) shoots. These calmodulins have electrophoretic mobilities on 10% (wt/vol) polyacrylamide gels at pH 8.3 in the presence of 6 M-urea and at pH 8.3 in the presence of 0.1% sodium dodecyl sulphate similar to that of bovine brain calmodulin. They interacted with rabbit skeletal-muscle troponin I in the presence of Ca2+. Barley and fungal calmodulins activated myosin light-chain kinase and phosphodiesterase in the presence of Ca2+, although the amounts needed were at least an order of magnitude greater than is required to produce the same effect with mammalian calmodulin. Amino acid analyses indicated a number of differences from the mammalian protein, most notably the absence of trimethyl-lysine. By using 125I-labeled calmodulin, a small amount of calmodulin-binding protein was detected in homogenates of barley and fungi. No protein corresponding to calmodulin could be found in Escherichia coli or yeast, although a relatively high concentration of a protein that bound calmodulin was detected in E. coli by this technique.