An ion-current mutant of Paramecium tetraurelia with defects in the primary structure and post-translational N-methylation of calmodulin

My work on pantophobiac A2 (pntA2), a behavioral mutant of Paramecium tetraurelia, suggest that the Ca++-binding protein calmodulin (CaM), and post-translation N-methylation of CaM, are important for Ca++-related ion-current function. Calmodulin from wild-type Paramecium has two sites of lysine-N-methylation. Both of these sites are almost fully methylated in vivo; thus wild-type calmodulin is a poor substrate for N-methylation in vitro. In contrast, pntA/2 CaM can be heavily N-methylated in vitro, suggesting that the mutant calmodulin is under-methylated in vivo. Amino-acid composition analysis showed that CaM lysine 115 is undermethylated in pntA2. Once pntA2 CaM is N-methylated, the [methyl-3H] group does not turn over in either wild-type or pntA2 cytoplasmic fractions. The methylating enzymes in pntA2 high-speed supernatant fractions are active, but may be less robust than those of the wild type, suggesting a possible control of these enzymes by CaM