Heteromeric three-stranded coiled coils designed using a Pb(II)(Cys) template mediated strategy.

Three-stranded coiled coils are peptide structures constructed from amphipathic heptad repeats. Here we show that it is possible to form pure heterotrimeric three-stranded coiled coils by combining three distinct characteristics: (1) a cysteine sulfur layer for metal coordination, (2) a thiophilic, trigonal pyramidal metalloid (Pb(II)) that binds to these sulfurs and (3) an adjacent layer of reduced steric bulk generating a cavity where water can hydrogen bond to the cysteine sulfur atoms. Cysteine substitution in an a site yields Pb(II)AB heterotrimers, while d sites provide pure Pb(II)CD or Pb(II)CD scaffolds.

Solution structure of a designed cyclic peptide ligand for nickel and copper ions.

Nuclear magnetic resonance (NMR) spectroscopy was used to study a cyclic peptide derived from the amino-terminal copper-and-nickel-binding (ATCUN) motif. The three-dimensional structure of the unliganded peptide in aqueous solution was solved by simulated annealing using distance constraints derived from Nuclear Overhauser Effects. A structural model for the Ni(II)-bound complex was also produced based on NMR evidence and prior spectroscopic data, which are consistent with crystal structures of linear ATCUN complexes.

Cysteinate protonation and water hydrogen bonding at the active-site of a nickel superoxide dismutase metallopeptide-based mimic: implications for the mechanism of superoxide reduction.

Nickel-containing superoxide dismutase (NiSOD) is a mononuclear cysteinate-ligated nickel metalloenzyme that catalyzes the disproportionation of superoxide into dioxygen and hydrogen peroxide by cycling between Ni(II) and Ni(III) oxidation states. All of the ligating residues to nickel are found within the first six residues from the N-terminus, which has prompted several research groups to generate NiSOD metallopeptide-based mimics derived from the first several residues of the NiSOD sequence. To assess the viability of using these metallopeptide-based mimics (NiSOD maquettes) to probe the mechanism of SOD catalysis facilitated by NiSOD, we computationally explored the initial step of the O2(-) reduction mechanism catalyzed by the NiSOD maquette {Ni(II)(SOD(m1))} (SOD(m1) = HCDLP CGVYD PA).

Metal-binding and redox properties of substituted linear and cyclic ATCUN motifs.

The amino-terminal copper and nickel binding (ATCUN) motif is a short peptide sequence found in human serum albumin and other proteins. Synthetic ATCUN-metal complexes have been used to oxidatively cleave proteins and DNA, cross-link proteins, and damage cancer cells. The ATCUN motif consists of a tripeptide that coordinates Cu(II) and Ni(II) ions in a square planar geometry, anchored by chelation sites at the N-terminal amine, histidine imidazole and two backbone amides.

Macrocyclization of the ATCUN motif controls metal binding and catalysis.

We report the design, synthesis, and characterization of macrocyclic analogues of the amino-terminal copper and nickel binding (ATCUN) motif. These macrocycles have altered pH transitions for metal binding, and unlike linear ATCUN motifs, the optimal cyclic peptide 1 binds Cu(II) selectively over Ni(II) at physiological pH. UV-vis and EPR spectroscopy showed that cyclic peptide 1 can coordinate Cu(II) or Ni(II) in a square planar geometry.

The importance of stereochemically active lone pairs for influencing Pb(II) and As(III) protein binding.

The toxicity of heavy metals, which is associated with the high affinity of the metals for thiolate rich proteins, constitutes a problem worldwide. However, despite this tremendous toxicity concern, the binding mode of As(III) and Pb(II) to proteins is poorly understood. To clarify the requirements for toxic metal binding to metalloregulatory sensor proteins such as As(III) in ArsR/ArsD and Pb(II) in PbrR or replacing Zn(II) in δ-aminolevulinc acid dehydratase (ALAD), we have employed computational and experimental methods examining the binding of these heavy metals to designed peptide models.

Pb-207 NMR spectroscopy reveals that Pb(II) coordinates with glutathione (GSH) and tris cysteine zinc finger proteins in a PbS3 coordination environment.

207Pb NMR spectroscopy can be used to monitor the binding of Pb(II) to thiol rich biological small molecules such as glutathione and to zinc finger proteins. The UV/visible (UV/Vis) absorption band centered at 334 nM and the observed 207Pb-signal in 207Pb NMR (δ~5750 ppm) indicate that glutathione binds Pb(II) in a trigonal pyramidal geometry (PbS3) at pH 7.5 or higher with a 1:3 molar ratio of Pb(II) to GSH.

Metallopeptide based mimics with substituted histidines approximate a key hydrogen bonding network in the metalloenzyme nickel superoxide dismutase.

Nickel superoxide dismutase (NiSOD) is a recently discovered superoxide dismutase that utilizes the Ni(III)/Ni(II) couple to facilitate the disproportionation of O(2)(*-) into H(2)O(2) and O(2). A key structural component of NiSOD is an elongated axial His-imidazole Ni(III) bond (2.3-2.

Probing variable axial ligation in nickel superoxide dismutase utilizing metallopeptide-based models: insight into the superoxide disproportionation mechanism.

Nickel superoxide dismutase (NiSOD) is a bacterial metalloenzyme that possesses a mononuclear Ni-center and catalyzes the disproportionation of O2*- by cycling between NiII and NiIII oxidation states. Herein we present evidence from several SOD active metallopeptide maquettes ([Ni(SODM2H(1)X)]; SODM2H(1)X = H2N-XCDLPCG-COOH; X = H, D, or A) that the Ni-center of NiSOD most likely remains five-coordinate during SOD catalysis using thin-film voltammetry. N3- and CN- titration studies suggest that O2*- disproportionation by [Ni(SODM2H(1)X)] proceeds via an outersphere mechanism.

The influence of amine/amide versus bisamide coordination in nickel superoxide dismutase.

Nickel superoxide dismutase (NiSOD) is a mononuclear nickel-containing metalloenzyme that catalyzes the disproportionation of superoxide by cycling between NiII and NiIII oxidation states. In the reduced NiII oxidation state, the metal center is ligated by two cysteinate sulfurs, one amide nitrogen, and one amine nitrogen (from the N-terminus), while in the oxidized NiIII state, an imidazole nitrogen coordinates to the metal center. Herein, we expand on a previous report in which we described a functional metallopeptide-based NiSOD model compound [NiII(SODM1)] (SODM1 = H2N-HCDLPCGVYDPA-COOH) by exploring how acylation of the N-terminus (producing [NiII(SODM1-Ac)]) influences the properties of the metallopeptide.