Cooperative Metal Binding and Helical Folding in Model Peptides of Treble‐Clef Zinc Fingers

Synergy in zinc fingers: The comparison between peptide folding and metal binding properties of two model peptides of treble‐clef zinc fingers presenting high affinities for zinc and cobalt reveals a cooperative effect: the metal folds the peptide into a α‐helix, which in turn strengthens the metal core. magnified image Two peptides, L_TC and ${{\rm L}{{{\rm T}\hfill \atop {\rm TC}\hfill}}}$ have been synthesised to model the treble‐clef zinc fingers encountered in many Zn(Cys)₄‐site‐containing proteins. Both are cyclic peptides with a linear tail grafted on a glutamate side chain of the cycle. They differ by the length of this tail, which lacks five amino acids in ${{\rm L}{{{\rm T}\hfill \atop {\rm TC}\hfill}}}$ compared to L_TC. Both peptides bind Zn²⁺ and Co²⁺ in 1:1 metal/peptide ratio and the structure of these complexes have been characterised by NMR, UV/Vis and CD spectroscopy. Both peptides fold the same way around the metal ion and they fully reproduce the classical fold of treble‐clef zinc fingers and display an extended hydrogen‐bond network around the coordinating sulfur atoms. The structures of the M⋅L_TC complexes reveal that the linear tail forms a short two‐turn α‐helix, present in the metallated form only. The formation of this helix constitutes a rare example of metal‐induced folding. The second turn of this helix is composed of the five amino acids that are absent in ${{\rm L}{{{\rm T}\hfill \atop {\rm TC}\hfill}}}$ . The study of the pH‐dependence of the Zn²⁺ binding constants shows that the metal ion is bound by four cysteinates above pH 5.2 and the binding constants are the highest reported so far. Interestingly, the binding constant of Zn⋅L_TC is about tenfold higher than that of Zn⋅${{\rm L}{{{\rm T}\hfill \atop {\rm TC}\hfill}}}$ . This difference clearly indicates that the helix, present in Zn⋅L_TC only, stabilises the Zn²⁺ complex by about 1.2 kcal mol⁻¹. The origin of this stabilisation is ascribed to an electrostatic interaction between the [ZnS₄]²⁻ centre and the helix. This reveals a cooperative effect: zinc binding allows the folding of the tail into a helix which, in turn, strengthens the zinc complex.