Characterization and purification of a soluble protein controlling Ca-channel activity in paramecium.
The analysis of the voltage-sensitive Ca++ channel of the unicellular eucaryote, Paramecium has been extended to a biochemical level based on recent observations that the transfer of cytoplasm from wild-type cells into mutants lacking Ca++-channel function ("pawn" in P. tetraurelia and "CNR" in P. caudatum) causes mutant cells to regain Ca++-channel function. We have microinjected various cytoplasmic fractions into mutant cells and measured the restored Ca++-channel function using a convenient behavioral assay. Following the "curing" activity, we characterized and purified the component from wild-type cytoplasm that can restore the function missing in cells carrying mutations in the cnrC gene. The curing factor is not an RNA, but a heat-labile, -SH-containing protein that appears to affect existing mutant channels on the ciliary membrane. We have purified this factor over 500-fold from the soluble cytoplasm using conventional techniques. The protein is of low apparent molecular weight (less than 30,000 daltons), acidic, soluble, and does not have the properties of calmodulin.
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000302:isolation & purification; D004553:Electric Conductivity; D007473:Ion Channels / Q000378:metabolism*; D008565:Membrane Proteins / Q000302:isolation & purification* / Q000378:metabolism; D008970:Molecular Weight; D009154:Mutation; D010247:Paramecium / Q000378:metabolism*; D015800:Protozoan Proteins
D002147:Calmodulin; C043358:CnrC protein, Paramecium; D007473:Ion Channels; D008565:Membrane Proteins; D015800:Protozoan Proteins; D002118:Calcium