Calmodulin Binding Proteins In Platelet Actomypsin

Platelet actomyosin (thrombosthenin) possesses a myosin-linked Ca2+ regulation and Ca2+ sensitivity is conferred to it by calmodulin through myosin light chain kinase. Calmodulin binding proteins if they are present in the actomyosin complex may have an important regulatory role in the contractile mechanism of platelet activation. To test this possibility an aceton powder was made from platelet actomyosin and extracted with an 8 M urea containing buffer. The extract was examined for the presence of calmodulin binding proteins by alkaline urea polyacrylamide gel electrophoresis. It was shown by this technique that some proteins in the actomyosin complex can form a Ca2+ dependent complex with both calmodulin and skeletal muscle troponin C (TNC is closely related to calmodulin) even in the presence of 8 M urea. Calmodulin binding proteins could be isolated from the extract by affinity chromatography in 8 M urea on TNC-Agarose column. 3 major proteins of 270 K, 6l K and 23 K molecular weight were eluted by EGTA and each of them was able to bind to calmodulin or TNC if Ca2+was present. At least one of these calmodulin binding proteins exerted a troponin I like effect when tested on reconstituted skeletal muscle actomyosin and the 23 K protein showed a close similarity to troponin I. the inhibitory protein of the actin linked Ca2+ regulatory system in skeletal muscle. It is presumed that calmodulin binding proteins may have a dual role in the regulation of platelet actomyosin. They can inhibit the Ca2+ dependent phosphorylation of myosin light chain and one or more of them may also exert an actin linked inhibitory effect.