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Taking into account the K D = 34.7 fM ( K D determined from competition with His in the present work) , K D 1 = 3.7 × 10 −9 M and K D 2 = 4.7 × 10 −7 M values for Cu(II)-His complexes at pH 7.4 (taken from 31 ) it can be estimated that approximately 0.7 nM of Cu(II) are bound to His mainly in the form of Cu•His 2 complex (Equations are presented in Materials and methods)