Cd(II) Capture Ability of an Immobilized, Fluorescent Hexapeptide

Abstract

Cys-Tyr-(YY), targeting Cd(II) ion sensing, was synthesized on various solid supports including two resins, as well as glass and quartz surfaces. The synthesis was based on the Fmoc (9-fluorenylmethoxycarbonyl) and the APTES (3-aminopropyltriethoxysilane) methodologies on the resin and silica supports, respectively. The immobilized ligand, except when coupled to a hydrophobic benzhydrylamine resin, showed a remarkably efficient, pHdependent Cd(II) capturing ability with a maximum binding capacity around neutral pH. The effect of contact time and metal ion concentration was also studied with a hydrophilic resin supported peptide (YY-NTG). The interaction of YY-NTG with Cd(II) was investigated by pH-potentiometric titrations in aqueous samples containing the resin beads and Cd(II). These studies, together with metal ion capturing experiments under buffer-controlled pH, prove that each immobilized peptide can bind one Cd(II) ion at pH=7.0 in the presence of one equivalent metal ion or metal ion excess. For Cd(II) binding to YY-NTG a notably high, K=1.310 10 apparent stability constant was determined (1:1 metal-to-ligand ratio, pH=7.0). Analytical results suggests that the concentration of Cd(II) can be measured below 200 nM with this silica-supported peptide. The usefulness of the probe was demonstrated by fluorescence spectroscopy.

Metal Exchange in the Interprotein ZnII‐Binding Site of the Rad50 Hook Domain: Structural Insights into CdII‐Induced DNA‐Repair Inhibition

“…The CI values of these Cd II hook complexes are two‐to‐four orders of magnitude more stable (CI=17–19) than the strongest ones with XPA zf (CI=12.8), CadC (CI=12.6), CmtR (CI=12.2), or MT2 (CI=14.4 for β‐domain and 15.8 for α‐domain) . Moreover, the complexes investigated here are much more stable than the Cd II complex with EDTA (CI=13.6), consensus CP1 zf (CCCC) zinc finger (CI=13.4), or short poly‐Cys peptides known to form highly stable complexes with Cd II , for instance Ac‐YCSSCY or Ac‐CC‐NH 2 , for which CI is 14.8 and 12.6, respectively . This brief overview clearly demonstrates that the zinc hook domain with substituted Cd II must exhibit unique stabilization effects that elevate thermodynamic stability.…”

“…RT,p HT,C C, and n.c. refer to reverset itration, pH titration,c ompetitionw ith metalc helator,a nd not calculated, respectively.C Iv aluesd erived from formation constantsp resented as 'not calculated' (n.c.) were determinedf rom published log b ijk data from potentiometric analyses. [38,39,62,63,83] Ligand [a] ReferenceM ethod of determination were not included in further discussion. Nonetheless, the results prove that Zn II -to-Cd II swap is af ast and efficient process that can occuri nside ac ell, causing Cd II -induced toxic effects.…”

“…[59] Moreover,the complexes investigated here are much more stable thant he Cd II complex with EDTA( CI = 13.6), [60] consensus CP1 zf (CCCC)z inc finger (CI = 13.4), [61] or short poly-Cys peptides known to form highly stable complexes with Cd II ,f or instance Ac-YCSSCY or Ac-CC-NH 2 ,f or which CI is 14.8 and 12.6, respectively. [62,63] This brief overview clearly demonstrates that the zinc hook domain with substituted Cd II muste xhibit unique stabilizatione ffects that elevatet hermodynamic stability. Molecular reasonsf or such stabilitya re discussed below.…”

Metalloproteinek fémkötőhelyein alapuló oligopeptidek, mint potenciális toxikus fémion érzékelők

“…A molekulák azonban egyértelmûen túl kicsik ahhoz, hogy a két fluorofór csoport a szabad peptidekben a Förster-távolságon kívül 41 Tapasztalataink és az irodalmi adatok áttekintése arra utalnak, hogy tényleges FRET effektuson alapuló oligopeptid receptorra épülõ fémion érzékelõt a fluorofór egységek fémion-kötõhelytõl távolabbi elhelyezésével lehetne kialakítani, és a molekulatervezésnél érdemes számítógépes modellszámításokra támaszkodni. 47 A CXXC szekvencia (ahol X bármilyen aminosavat jelöl) a metallotioneninek mellett fémion szállító-, raktározó-, ill. dajkafehérjékben egyaránt elõfordul. Egyebek mellett a Hg 2+ -kötõ MerP, 48 a bakteriális vagy humán ATP7A és ATP7B fehérjék, 49 ill. a réz-chaperon bakteriális Atx1, vagy az ezzel analóg humán Atox1 (HAH1) 50 is ezt a szekvenciát alkalmazza "szoft" karakterû fémionok megkötésére.…”

“…47 A titrálások révén meghatározott protonálódási és komplexképzõdési állandók jó összhangban vannak a Cd 2+ -megkötõdés vizsgálatok eredményeivel. A legtöbb kísérletnél használt Cd 2+ :YY-NTG 0,66:1 arányra, illetve az ilyen összetételû minták koncentrációira szimulált eloszlásgörbe (12.…”