Localization of hydrophobic sites in calmodulin and skeletal muscle troponin C studied using tryptic fragments: a simple method of their preparation.
The exposure of hydrophobic sites on calmodulin, skeletal muscle troponin C and their tryptic fragments was investigated using Phenyl-Sepharose chromatography. A strong binding of both proteins and their fragments corresponding to the NH2-terminal halves of polypeptide chain of respective proteins in the presence of calcium ions was observed. Only a weak interaction with Phenyl-Sepharose or its lack was observed under these conditions for fragments corresponding to the COOH-terminal halves of calmodulin and troponin C, respectively. The elution of the samples from Phenyl-Sepharose column with ethylene glycol gradient allowed to compare relative hydrophobicity of both proteins and their fragments. The results show that hydrophobic properties of calmodulin and troponin C are virtually preserved in their fragments obtained as a result of their cleavage by trypsin in half. They also indicated that the exposure of hydrophobic residues caused by the binding of calcium ions takes place mainly in the NH2-terminal halves of polypeptide chains of both proteins. A simple method of purification of tryptic fragments of both proteins based on the difference in the strength of their interactions with Phenyl-Sepharose is described.
D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism*; D002846:Chromatography, Affinity; D009124:Muscle Proteins / Q000378:metabolism*; D009132:Muscles / Q000378:metabolism*; D010446:Peptide Fragments / Q000302:isolation & purification* / Q000378:metabolism; D011485:Protein Binding; D012685:Sepharose / Q000031:analogs & derivatives; D014336:Troponin / Q000378:metabolism*; D019209:Troponin C; D014357:Trypsin
D002135:Calcium-Binding Proteins; D002147:Calmodulin; D009124:Muscle Proteins; D010446:Peptide Fragments; D014336:Troponin; D019209:Troponin C; C020927:phenyl-sepharose; D012685:Sepharose; D014357:Trypsin; D002118:Calcium