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Several pH-dependent low-spin ferric haem forms are identified in a frozen solution of the ferric 121Cys→Ser mutant of Drosophila melanogasterhaemoglobin (DmHb1*) using electron paramagnetic resonance (EPR) techniques. Different forms with EPR parameters typical of bis-histidine coordinated haem iron centers were observed. Strong pH-dependent changes in the EPR signatures were observed related to changes in the haem pocket. The pulsed EPR data indicate that both the distal and proximal histidine exhibit a large libration around the Fe–NHis axis. The resonance Raman spectra of the CO-ligated ferrous form of Drosophila melanogasterhaemoglobin are typical of an open conformation, with little stabilization of the CO ligand by the surrounding amino-acid residues. The EPR data of the cyanide-ligated ferric DmHb1* indicates a close similarity with cyanide-ligated ferric myoglobin. The structural characteristics of DmHb1* are found to clearly differ from those of other bis-histidine-coordinated globins.

Neisseria gonorrhoeae has the capacity to acquire iron from its human host by removing this essential nutrient from serum transferrin. The transferrin binding proteins, TbpA and TbpB, constitute the outer membrane receptor complex responsible for binding transferrin, extracting the tightly bound iron from the host-derived molecule, and transporting iron into the periplasmic space of this Gram-negative bacterium. Once iron is transported across the outer membrane, ferric binding protein A (FbpA) moves the iron across the periplasmic space and initiates the process of transport into the bacterial cytosol. The results of the studies reported here define the multiple steps in the iron transport process in which TbpA and TbpB participate. Using the SUPREX technique for assessing the thermodynamic stability of protein–ligand complexes, we report herein the first direct measurement of periplasmic FbpA binding to the outer membrane protein TbpA. We also show that TbpA discriminates between apo- and holo-FbpA; i.e. the TbpA interaction with apo-FbpA is higher affinity than the TbpA interaction with holo-FbpA. Further, we demonstrate that both TbpA and TbpB individually can deferrate transferrin without energy supplied from TonB resulting in sequestration by apo-FbpA.

Metal chelates with biomolecules are increasingly used in animal supplementation to increase the bioavailability of essential trace elements. However, the transfer of the chelates is not well understood and speciation studies may bring a comprehensive insight to further investigate the biological uptake mechanism(s) implicated. An analytical method was developed for the characterization of the water-soluble metal complexes in animal feed supplements obtained by reaction of a metal salt with a non-GMO soybean enzymatic digest. The method was based on fractionation of the extract by size-exclusion chromatography followed by the analysis of the metal-containing fraction by reversed-phase nanoHPLC with parallel ICP MS and electrospray MS/MS detection. The metal complexes were identified in the mass spectra owing to the Cu characteristic isotopic pattern; the complexation was corroborated by the presence of a peak corresponding to the non-metallated peptide. The study demonstrated the feasibility of SEC-ICP MS to produce characteristic metal (Cu, Zn, Mn, Fe) distribution patterns, which can be of interest to test batch-to-batch reproducibility and to determine the origin of the supplement. The use of the method could be extended to animal feeds prepared using the metal-chelated complexes. Electrospray MS/MS allowed the identification of a number of Cu complexes with peptides. Four different structure conformations were modeled by means of molecular mechanics investigations to assess the chelation stability.

Data on the metal-binding behaviour of circa 20 recombinant metallothioneins (MTs) from evolutionary divergent organisms, gathered after years of systematic research, are here comprehensively analyzed. The consideration of four independent in vivo and in vitro metal-binding features reveals a gradation of the metal-binding character of the MTs considered that significantly coincides in a robust new classification: a stepwise gradation between Zn- and Cu-thioneins. The intermediate positions in this list are occupied by a group of polyvalent MTs, exhibiting a merging Zn-/Cu-thionein character that would suit general metal handling purposes. In contrast, the extreme positions are respectively occupied by those MTs that would have evolved to fulfil specialized Zn- or Cu-related physiological roles. Overall, the analyzed trends allow the proposal of a chemically- and biologically-sound new reflection on MT classification criteria.

Although carcinogenic metals have been known to disrupt a wide range of cellular processes the precise mechanism by which they exert their carcinogenic effects is not known. Over the last decade or two, studies in the field of metal carcinogenesis suggest that epigenetic mechanisms may play a role in metal-induced carcinogenesis. In this review we summarize the evidence demonstrating that exposure to carcinogenic metals such as nickel, arsenic, chromium, and cadmium can perturb DNA methylation levels as well as global and gene specific histone tail posttranslational modification marks. We also wish to emphasize the importance of understanding that gene expression can be regulated by both genetic and epigenetic mechanisms and both these must be considered when studying the mechanism underlying the toxicity and cell transforming ability of carcinogenic metals and other toxicants, as well as aberrant changes in gene expression that occur during disease states such as cancer.

Urease, the first enzyme to be crystallized, contains a dinuclear nickel metallocenter that catalyzes the decomposition of urea to produce ammonia, a reaction of great agricultural and medical importance. Several mechanisms of urease catalysis have been proposed on the basis of enzyme crystal structures, model complexes, and computational efforts, but the precise steps in catalysis and the requirement of nickel versus other metals remain unclear. Purified bacterial urease is partially activated via incubation with carbon dioxide plus nickel ions; however, in vitro activation also has been achieved with manganese and cobalt. In vivo activation of most ureases requires accessory proteins that function as nickel metallochaperones and GTP-dependent molecular chaperones or play other roles in the maturation process. In addition, some microorganisms control their levels of urease by metal ion-dependent regulatory mechanisms.

Wilson’s disease, an autosomal recessive disease of copper accumulation and copper toxicity primarily in the liver and brain, has been the engine that has driven the development of anticopper drugs. Here we first briefly review Wilson’s disease, then review the four anticopper drugs used to treat Wilson’s disease. We then discuss the results of therapy with anticopper drugs in Wilson’s disease, with special emphasis on the newer and better drugs, zinc and tetrathiomolybdate. We then discuss new areas of anticopper therapy, lowering copper availability with tetrathiomolybdate as a therapy in fibrotic, inflammatory, and autoimmune disorders. Many of the cytokines which promote these disorders are copper dependent, and lowering copper availability lessens the activity of these cytokines, favorably influencing a variety of disease processes. Copper in the blood can be thought of as in two pools. One pool is covalently bound in ceruloplasmin, a protein containing six coppers, synthesized by the liver and secreted into the blood. Ceruloplasmin copper accounts for almost 85 to 90% of the blood copper in normal people. This copper is tightly bound and not readily available for cellular uptake and copper toxicity. The other 10–15% of copper is more loosely bound to albumin and other small molecules in the blood, and is readily and freely available to cells and available to cause copper toxicity, if this pool of copper is increased. We call this latter pool of copper “free” copper because of its more ready availability. However, it should be understood that it is not completely free, always being bound to albumin and other molecules. It is this pool of free copper that is greatly expanded in untreated Wilson’s patients undergoing copper toxicity.

Gallium-based anticancer chemotherapeutics are appreciably progressing in clinical studies. A steady interest of drug developers and clinicians in gallium compounds is due to a proven ability of gallium cations to inhibit tumour growth, on the one hand, and enhanced bioavailability and moderate toxicity provided by the conversion of gallium into chelate complexes, on the other. One of the complexes suitable for a more convenient oral administration is tris(8-quinolinolato)gallium(III) (KP46). Nominated from a range of gallium complexes for the clinical stage of development, KP46 has finished phase I trials with the outcome of promising tolerability and evidence of clinical activity in renal cell carcinoma. Therefore, there is obviously a need to codify and critically evaluate the continuing advances in the emergence of KP46 as a lead-drug candidate. Additionally, many questions remain unanswered regarding the relevant biological reactivity, modes of delivery and action and potential cell target(s) of KP46. The timely publication of the present review is also an attempt to shed light on these pertinent drug assets and to accelerate research activities towards further clinical development of KP46.

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