Calmodulin and troponin C: affinity chromatographic study of divalent cation requirements for troponin I inhibitory peptide (residues 104-115), mastoparan and fluphenazine binding.
The different conformations induced by the binding of Mg2+ or Ca2+ to troponin C (TnC) and calmodulin (CaM) results in the exposure of various interfaces with potential to bind target compounds. The interaction of TnC or CaM with three affinity columns with ligands of either the synthetic peptide of troponin I (TnI) inhibitory region (residues 104-115), mastoparan (a wasp venom peptide), or fluphenazine (a phenothiazine drug) were investigated in the presence of Mg2+ or Ca2+. TnC and CaM in the presence of either Ca2+ or Mg2+ bound to the TnI peptide 104-115. The cation specificity for this interaction firmly establishes that the TnI inhibitory region binds to the high affinity sites of TnC (most likely the N-terminal helix of site III) and presumably the homologous region of CaM. Mastoparan interacted strongly with both proteins in the presence of Ca2+ but, in the presence of Mg2+, did not bind to TnC and only bound weakly to CaM. Fluphenazine bound to TnC and CaM only in the presence of Ca2+. When the ligands interacted with either proteins there was an increase in cation affinity, such that TnC and CaM were eluted from the TnI peptide or mastoparan affinity column with 0.1 M EDTA compared with the 0.01 M EDTA required to elute the proteins from the fluphenazine column. The interaction of these ligands with their receptor sites on TnC and CaM require a specific and spatially correct alignment of hydrophobic and negatively charged residues on these proteins.(ABSTRACT TRUNCATED AT 250 WORDS)
Department of Biochemistry, University of Alberta, Edmonton, Canada.
J E
JE
Van Eyk
R S
RS
Hodges
D001514:Bee Venoms / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism*; D002413:Cations, Divalent / Q000378:metabolism*; D002846:Chromatography, Affinity; D005476:Fluphenazine / Q000378:metabolism*; D066298:In Vitro Techniques; D036341:Intercellular Signaling Peptides and Proteins; D010455:Peptides; D014336:Troponin / Q000378:metabolism*; D019209:Troponin C; D019210:Troponin I; D014862:Wasp Venoms / Q000378:metabolism*
D001514:Bee Venoms; D002147:Calmodulin; D002413:Cations, Divalent; D036341:Intercellular Signaling Peptides and Proteins; D010455:Peptides; D014336:Troponin; D019209:Troponin C; D019210:Troponin I; D014862:Wasp Venoms; C037500:mastoparan; D005476:Fluphenazine