title: The characterization for the binding of calcium and terbium to Euplotes octocarinatus centrin.
abstract: Centrin is a member of the EF-hand superfamily that plays critical role in the centrosome duplication and separation. In the present paper, we characterized properties of metal ions binding to Euplotes octocarinatus centrin (EoCen) by fluorescence spectra and circular dichroism (CD) spectra. Changes of fluorescence spectra and alpha-helix contents of EoCen proved that Tb(3+) and Ca(2+) induced great conformational changes of EoCen resulting in exposing hydrophobic surfaces. At pH 7.4, Ca(2+) (and Tb(3+)) bond with EoCen at the ratio of 4:1. Equilibrium experiment indicated that Ca(2+) and Tb(3+) exhibited different binding capabilities for C- and N-terminal domains of protein. C-terminal domain bond with Ca(2+) or Tb(3+) approximately 100-fold more strongly than N-terminal. Aromatic residue-sensitized Tb(3+) energy transfer suggested that site IV bond to Tb(3+) or Ca(2+) more strongly than site III. Based on fluorescence titration curves, we reckoned the conditional binding constants of EoCen site IV quantitatively to be K(IV)=(1.23+/-0.51)x10(8)M(-1) and K(IV)=(6.82+/-0.33)x10(5)M(-1) with Tb(3+) and Ca(2+), respectively. Metal ions bond to EoCen in the order of IV>III>II, I.
authors.affiliation: Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education, Institute of Molecular Science, Shanxi University, Taiyuan 030006, China.
authors.forename: Zhao
authors.initials: Z
authors.lastname: Yaqin
authors.forename: Feng
authors.initials: F
authors.lastname: Jiuying
authors.forename: Liang
authors.initials: L
authors.lastname: Aihua
authors.forename: Yang
authors.initials: Y
authors.lastname: Binsheng