Calcium-induced exposure of a hydrophobic surface on calmodulin

Biochemistry. 1980 Aug 5;19(16):3814-9. doi: 10.1021/bi00557a025.


Interactions between calmodulin (CaM) and several hydrophobic fluorescent probes were characterized in order to determine if CaM expresses hydrophobic binding sites in the presence of Ca2+. Several classes of fluorescent probes capable of sensing exposure of hydrophobic binding sites on proteins were found to bind to CaM, and these interactions were greatly enhanced by Ca2+. In the presence of Ca2+, the fluorescence intensity of 9-anthroylcholine (9AC) was increased 24-fold by CaM, with a shift in the fluorescence emission maximum from 514 to 486 nm. The fluorescence intensity of 8-anilino-1-naphthalenesulfonate (Ans) was enhanced 27-fold with an emission maximum shift from 540 to 488 nm in the presence of CaM and Ca2+. Similar results were obtained with the uncharged fluorescent ligand, N-phyenyl-1-naphthylamine. With all three fluorescent dyes, the fluorescence changes caused by CaM in the absence of Ca2+ were minor compared to those observed with CaM and Ca2+. Direct binding studies using equilibrium dialysis demonstrated that CaM can bind four to six molecules of 9AC or two to three molecules of Ans in a calcium-dependent manner. The effects of various amphiphilic compounds on the Ca2+-dependent complex formation between CaM and the Ca2+-sensitive phosphodiesterase or troponin I were investigated. Trifluoperazine (TFP) and 9AC inhibited CaM stimulation of the Ca2+-sensitive phosphodiesterase. The Ca2+-dependent binding of the phosphodiesterase to CaM-Sepharose was also inhibited by TFP, 9AC, and Ans. Furthermore, binding of CaM to troponin I-Sepharose was inhibited by these ligands. Consistent with these data was the observation that troponin I antagonized binding of 9AC to CaM. These data indicate that binding of Ca2+ to CaM results in exposure of a domain with considerable hydrophobic character, and binding of hydrophobic ligands to this domain antagonizes CaM-protein interactions. It is proposed that this hydrophobic domain may serve as the interface for the Ca2+-dependent binding of CaM to the phosphodiesterase or troponin I.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Binding Sites
  • Calcium*
  • Calcium-Binding Proteins*
  • Calmodulin*
  • Cattle
  • Egtazic Acid
  • Fluorescent Dyes
  • Myocardium / enzymology
  • Phosphoric Diester Hydrolases / metabolism
  • Protein Binding
  • Protein Conformation
  • Spectrometry, Fluorescence
  • Troponin


  • Calcium-Binding Proteins
  • Calmodulin
  • Fluorescent Dyes
  • Troponin
  • Egtazic Acid
  • Phosphoric Diester Hydrolases
  • Calcium