Sarah L. J. Michel

Functional solitare- and trans-hybrids, the synthesis, characterization, electrochemistry and reactivity of porphyrazine/phthalocyanine hybrids bearing nitro and amino functionality

We report here for the first time a new and improved approach to the complete synthesis and characterization of the unsymmetrical metallo- and metal-free phthalocyanine/porphyrazine (Pz/Pc) hybrids bearing peripheral nitro or amine functionality, H2[Pz(n-Pr)6(BzNO2)] or H2[Pz(n-Pr)4(BzNO2)2], where Bz = benzo, n-Pr = n-propyl, fused on the one or two benzenoid ring which form from the reaction of two different precursors, 3,4-bis-(n-propyl)-pyroline-2,5-diimine and 6/7-nitro-1,3,3-tricholoro isoindoline, with the base-catalyzed direct condensation method. The direct condensation method used in this study enable us to skip magnesium porphyrazine (MgPz) stage and limits the formation of undesired hybrid compounds when compared with cross-condensation method. Centrally metallated solitare- and trans-Pz/Pc hybrids, M[Pz(n-Pr)6(BzNO2)] or M[Pz(n-Pr)4(BzNO2)2](M = Zn(II) or Mn(II)), synthesized in this study are somewhat more soluble than the free ones. Metallo Pz/Pc hybrids were prepared by treatment of free-soitare and trans hybrids with the appropriate metal salts in anhydrous solvent mixtures. We observed that the solubility of solitare- and/or trans-Pz/Pc hybrids increases after nitro-substituted Pz/Pc hybrids are reduced to amine substituted ones. The reactivity of the NH2 group of the solitare-Zn[Pz(n-Pr)6(BzNH2)] was tested by the condensation with ferrocenecarboxyaldehyde resulting in the formation of the ferrocene-substituted hybrid. The newly synthesized compounds have been characterized by elemental analaysis, FT IR, 1H NMR, UV-vis and MS spectroscopy. Electrochemical measurements of the complexes characterized in this study showed that the attachment of NO2 and NH2 moieties at the periphery makes a monitorable effect on redox properties of the macrocycle and metal center.

Holo-Ni(II)HpNikR is an asymmetric tetramer containing two different nickel binding sites

The metalloregulatory protein NikR from Helicobacter pylori (HpNikR) is a master regulator of gene expression which both activates and represses specific genes in response to nickel availability. Here, we report the first crystal structure (at 2.37 Å resolution) of Ni(II)HpNikR prepared directly from the holo protein. The protein contains four nickel ions located in two distinct coordination environments. Two nickel ions are bound to sites in a four-coordinate square-planar geometry as predicted on the basis of the structures of NikR from Escherichia coli and Pyrococcus horikoshii . The remaining two nickel ions are bound to sites with unexpected 5- or 6-coordination geometries which were previously thought to be involved in nickel incorporation into the protein. The nickel with 5-/6-coordination geometry utilizes three histidines from two separate monomeric HpNikR units along with two or three water molecules as ligands. The spatial location of the nickel in the 5-/6-coordinate site is within approximately 5 Å of the expected site if a 4-coordinate square-planar geometry occurred. Two of the histidines that participate as ligands in the 5-/6-coordinate site would also participate as ligands if the 4-coordinate site was occupied, making it impossible for both sites to be occupied simultaneously. DFT calculations show that the 5-/6-coordinate geometries are energetically favorable when the local protein environment is included in the calculations. The presence of two distinct coordination environments in HpNikR is suggested to be related to the specificity and binding affinity of this transcription factor for DNA.

Cysteine Oxidation Enhanced by Iron in Tristetraprolin, A Zinc Finger Peptide

Tristetraprolin (TTP or NUP475) is a non-classical zinc finger protein that is involved in inflammatory response. TTP regulates the production of cytokines by binding to specific mRNA sequences. TTP contains two Cys(3)His metal binding domains that can coordinate zinc, cobalt, ferric and ferrous iron. When zinc, cobalt, ferric or ferrous iron are bound, TTP peptides can bind to their cognate RNA. During inflammation there are increased levels of reactive oxygen species and iron. It has been proposed that reactive oxygen species may play a role in regulating zinc finger protein function by oxidizing cysteine thiolates that bind zinc and inactivating the protein. To elucidate the effect of the reactive oxygen species H(2)O(2) on the integrity of TTP and its ability to bind to target RNA, a simple and rapid assay using cobalt as a spectroscopic probe for zinc was developed. The oxidative susceptibility of peptides consisting of the zinc binding domains of a single zinc finger domain of TTP, TTP-D1 and the tandem zinc finger domains of TTP, TTP-2D was measured. Fe(II)-TTP-D1 and Fe(II)-TTP-2D were more rapidly oxidized by H(2)O(2) than their Zn(II) bound counterparts. Electron paramagnetic resonance (EPR) spin trapping using 2-ethoxycarbonyl-2-methyl-3,4-dihydro-2H-pyrrole-1-oxide (EMPO) demonstrated that oxidation of ferrous iron substituted TTP-D1 and TTP-2D resulted in the formation of hydroxyl radicals via Fenton chemistry. The oxidized peptides exhibited a diminished affinity for target RNA compared to their unoxidized counterparts suggesting that oxidation of TTP inactivates the protein.

Switching Metal Ion Coordination and DNA Recognition in a Tandem CCHHC-type Zinc Finger Peptide

Neural Zinc Finger Factor-1 (NZF-1) and Myelin Transcription Factor 1 (MyT1) are two homologous nonclassical zinc finger (ZF) proteins that are involved in the development of the central nervous system (CNS). Both NZF-1 and MyT1 contain multiple ZF domains, each of which contains an absolutely conserved Cys2His2Cys motif. All three cysteines and the second histidine have been shown to coordinate Zn(II); however, the role of the first histidine remains unresolved. Using a functional form of NZF-1 that contains two ZF domains (NZF-1-F2F3), mutant proteins in which each histidine was sequentially mutated to a phenylalanine were prepared to determine the role(s) of the histidine residues in DNA recognition. When the first histidine is mutated, the protein binds Zn(II) in an analogous manner to the native protein. Surprisingly, this mutant does not bind to target DNA (β-RAR), suggesting that the noncoordinating histidine is critical for sequence selective DNA recognition. The first histidine will coordinate Zn(II) when the second histidine is mutated; however, the overall fold of the protein is perturbed leading to abrogation of DNA binding. NZF-1-F2F3 selectively binds to a specific DNA target sequence (from β-RAR) with high affinity (nM); while its homologue MyT1 (MyT1-F2F3), which is 92% identical to NZF-1-F2F3, binds to this same DNA sequence nonspecifically. A single, nonconserved amino acid residue in NZF-1-F2F3 is shown to be responsible for this high affinity DNA binding to β-RAR. When this residue (arginine) is engineered into the MyT1-F2F3 sequence, the affinity for β-RAR DNA increases.

Lanthanide porphyrazine sandwich complexes: synthetic, structural and spectroscopic investigations

The synthesis of four novel lanthanide tetraazaporphyrin (porphyrazine) sandwich complexes is reported: [Ce(OPTAP)2] (2), [Lu(OPTAP)2] (3), [Eu(OPTAP)2] (4) and [Eu2(OPTAP)3] (5) (OPTAP = 2,3,7,8,12,13,17,18-octapropyltetraazaporphyrinato). The structure of 2 has been unequivocally established by an X-ray crystallographic study. The π radical nature of 3 and 4 has been confirmed by UV-vis, IR, and EPR spectroscopy. Cyclic voltammetry results show that 2–5 are more easily oxidized than 1 (H2OPTAP) by 0.41, 0.56, 0.48 and 0.40 V respectively.