Copper Chaperone Research Articles

Copper chaperone antioxidant 1: multiple roles and a potential therapeutic target.

Copper (Cu) was recently demonstrated to play a critical role in cellular physiological and biochemical processes, including energy production and maintenance, antioxidation and enzymatic activity, and signal transduction. Antioxidant 1 (ATOX1), a chaperone of Cu previously named human ATX1 homologue (HAH1), has been found to play an indispensable role in maintaining cellular Cu homeostasis, antioxidative stress, and transcriptional regulation. In the past decade, it has also been found to be involved in a variety of diseases, including numerous neurodegenerative diseases, cancers, and metabolic diseases. Recently, increasing evidence has revealed that ATOX1 is involved in the regulation of cell migration, proliferation, autophagy, DNA damage repair (DDR), and death, as well as in organism development and reproduction. This review summarizes recent advances in the research on the diverse physiological and cytological functions of ATOX1 and the underlying mechanisms of its action in human health and diseases. The potential of ATOX1 as a therapeutic target is also discussed. This review aims to pose unanswered questions related to ATOX1 biology and explore the potential use of ATOX1 as a therapeutic target.

Ceria Nanoparticles as Copper Chaperones that Activate SOD1 for Synergistic Antioxidant Therapy to Treat Ischemic Vascular Diseases (Adv. Mater. 16/2023)

Therapeutic Antioxidants In article number 2208989, Gwang-Bum Im, Young Geon Kim, Tae Yong Yoo, Taeghwan Hyeon, Suk Ho Bhang, and co-workers report the therapeutic effect of copper-deposited ceria nanoparticles (CuCe NPs) on ischemic vascular diseases. They exhibit enhanced antioxidant effects over pristine ceria nanoparticles as the released copper buffers the depletion of glutathione while providing the bioavailable copper as a cofactor for the antioxidant enzyme, superoxide dismutase 1.

Systematic pan-cancer analysis identifies SLC31A1 as a biomarker in multiple tumor types

BackgroundSolute Carrier Family 31 Member 1 (SLC31A1) has recently been identified as a cuproptosis-regulatory gene. Recent studies have indicated that SLC31A1 may play a role in colorectal and lung cancer tumorigenesis. However, the role of SLC31A1 and its cuproptosis-regulatory functions in multiple tumor types remains to be further elucidated.MethodsOnline websites and datasets such as HPA, TIMER2, GEPIA, OncoVar, and cProSite were used to extract data on SLC31A1 in multiple cancers. DAVID and BioGRID were used to conduct functional analysis and construct the protein–protein interaction (PPI) network, respectively. The protein expression data of SLC31A1 was obtained from the cProSite database.ResultsThe Cancer Genome Atlas (TCGA) datasets showed increased SLC31A1 expression in tumor tissues compared with non-tumor tissues in most tumor types. In patients with tumor types including adrenocortical carcinoma, low-grade glioma, or mesothelioma, higher SLC31A1 expression was associated with shorter overall survival and disease-free survival. S105Y was the most prevalent point mutation in SLC31A1 in TCGA pan-cancer datasets. Moreover, SLC31A1 expression was positively correlated with the infiltration of immune cells such as macrophages and neutrophils in tumor tissues in several tumor types. Functional enrichment analysis showed that SLC31A1 co-expressed genes were involved in protein binding, integral components of the membrane, metabolic pathways, protein processing, and endoplasmic reticulum. Copper Chaperone For Superoxide Dismutase, Phosphatidylinositol-4,5-Bisphosphate 3-Kinase Catalytic Subunit Alpha and Solute Carrier Family 31 Member 2 were copper homeostasis-regulated genes shown in the PPI network, and their expression was positively correlated with SLC31A1. Analysis showed there was a correlation between SLC31A1 protein and mRNA in various tumors.ConclusionsThese findings demonstrated that SLC31A1 is associated with multiple tumor types and disease prognosis. SLC31A1 may be a potential key biomarker and therapeutic target in cancers.

The Sensory Histidine Kinase CusS of Escherichia coli Senses Periplasmic Copper Ions.

Two-component regulatory systems composed of a membrane-bound sensor/sensory histidine kinase (HK) and a cytoplasmic, DNA-binding response regulator (RR) are often associated with transenvelope efflux systems, which export transition metal cations from the periplasm directly out of the cell. Although much work has been done in this field, more evidence is needed for the hypothesis that the respective two-component regulatory systems are indeed sensing periplasmic ions. If so, a regulatory circuit between the concentration of periplasmic metal cations, sensing of these metals, and control of expression of the genes for transenvelope efflux systems that remove periplasmic cations can be assumed. Escherichia coli possesses only one transenvelope efflux system for metal cations, the Cus system for export of Cu(I) and Ag(I). It is composed of the transenvelope efflux system CusCBA, the periplasmic copper chaperone CusF, and the two-component regulatory system CusS (HK) and CusR (RR). Using phoA- and lacZ-reporter gene fusions, it was verified that an assumed periplasmic part of CusS is located in the periplasm. CusS was more important for copper resistance in E. coli under anaerobic conditions than under aerobic conditions and in complex medium more than in mineral salts medium. Predicted copper-binding sites in the periplasmic part of CusS were identified that, individually, were not essential for copper resistance but were in combination. In summary, evidence was obtained that the two-component regulatory system CusSR that controls expression of cusF and cusCBA does indeed sense periplasmic copper ions. IMPORTANCE Homeostasis of essential-but-toxic transition metal cations such as Zn(II) and Cu(II)/Cu(I) is an important contributor to the fitness of environmental bacteria and pathogenic bacteria during their confrontation with an infected host. Highly efficient removal of threatening concentrations of these metals can be achieved by the combined actions of an inner membrane with a transenvelope efflux system, which removes periplasmic ions after their export from the cytoplasm to this compartment. To understand the resulting metal cation homeostasis in the periplasm, it is important to know if a regulatory circuit exists between periplasmic metal cations, their sensing, and the subsequent control of the expression of the transenvelope efflux system. This publication adds evidence to the hypothesis that two-component regulatory systems in control of the expression of genes for transenvelope efflux systems do indeed sense metal cations in the periplasm.

Ceria Nanoparticles as Copper Chaperones that Activate SOD1 for Synergistic Antioxidant Therapy to Treat Ischemic Vascular Diseases.

All exogenous nanomaterials undergo rapid biotransformation once injected into the body and fall short of executing the intended purpose. Here,it is reported that copper-deposited ceria nanoparticles (CuCe NPs) exhibit enhanced antioxidant effects overpristine ceria nanoparticles, as the released copper buffers the depletion of glutathione while providing the bioavailable copper as a cofactor for the antioxidant enzyme, superoxide dismutase 1. The upregulated intracellular antioxidants along with the ceria nanoparticles synergistically scavenge reactive oxygen species and promote anti-inflammation and M2 polarization of macrophages by modulating signal transducer and activator of transcription 1 and 6 (STAT1 and STAT6). The therapeutic effect of CuCe NPs is demonstrated in ischemic vascular diseases (i.e., murine models of hindlimb ischemia and myocardial infarction) in which the copper-deposition affords increased perfusion and alleviation in tissue damage. The results provide rationale that metal oxide nanomaterials can be designed in a way to induce the upregulation of specific biological factors for optimal therapeutic performance.

Trace copper-mediated asexual development via a superoxide dismutase and induction of AobrlA in Aspergillus oryzae.

The filamentous fungus Aspergillus oryzae, in which sexual reproduction remains to be discovered, proliferates mainly via asexual spores (conidia). Therefore, despite its industrial importance in food fermentation and recombinant protein production, breeding beneficial strains by genetic crosses is difficult. In Aspergillus flavus, which is genetically close to A. oryzae, structures known as sclerotia are formed asexually, but they are also related to sexual development. Sclerotia are observed in some A. oryzae strains, although no sclerotia formation has been reported in most strains. A better understanding of the regulatory mechanisms underlying sclerotia formation in A. oryzae may contribute to discover its sexual development. Some factors involved in sclerotia formation have been previously identified, but their regulatory mechanisms have not been well studied in A. oryzae. In this study, we found that copper strongly inhibited sclerotia formation and induced conidiation. Deletion of AobrlA encoding a core regulator of conidiation and ecdR involved in transcriptional induction of AobrlA suppressed the copper-mediated inhibition of sclerotia formation, suggesting that AobrlA induction in response to copper leads not only to conidiation but also to inhibition of sclerotia formation. In addition, deletion of the copper-dependent superoxide dismutase (SOD) gene and its copper chaperone gene partially suppressed such copper-mediated induction of conidiation and inhibition of sclerotia formation, indicating that copper regulates asexual development via the copper-dependent SOD. Taken together, our results demonstrate that copper regulates asexual development, such as sclerotia formation and conidiation, via the copper-dependent SOD and transcriptional induction of AobrlA in A. oryzae.

Genome-Wide Identification and Characterization of Copper Chaperone for Superoxide Dismutase (CCS) Gene Family in Response to Abiotic Stress in Soybean

Copper Chaperone For Superoxide Dismutase (CCS) genes encode copper chaperone for Superoxide dismutase (SOD) and dramatically affect the activity of SOD through regulating copper delivery from target to SOD. SOD is the effective component of the antioxidant defense system in plant cells to reduce oxidative damage by eliminating Reactive oxygen species (ROS), which are produced during abiotic stress. CCS might play an important role in abiotic stress to eliminate the damage caused by ROS, however, little is known about CCS in soybean in abiotic stress regulation. In this study, 31 GmCCS gene family members were identified from soybean genome. These genes were classified into 4 subfamilies in the phylogenetic tree. Characteristics of 31 GmCCS genes including gene structure, chromosomal location, collinearity, conserved domain, protein motif, cis-elements, and tissue expression profiling were systematically analyzed. RT-qPCR was used to analyze the expression of 31 GmCCS under abiotic stress, and the results showed that 5 GmCCS genes(GmCCS5, GmCCS7, GmCCS8, GmCCS11 and GmCCS24) were significantly induced by some kind of abiotic stress. The functions of these GmCCS genes in abiotic stress were tested using yeast expression system and soybean hairy roots. The results showed that GmCCS7/GmCCS24 participated in drought stress regulation. Soybean hairy roots expressing GmCCS7/GmCCS24 showed improved drought stress tolerance, with increased SOD and other antioxidant enzyme activities. The results of this study provide reference value in-depth study CCS gene family, and important gene resources for the genetic improvement of soybean drought stress tolerance.

The stepwise dissociation of the Zn(II)-bound Atox1 homodimer and its energetic asymmetry

The copper chaperone Atox1, which is featured by a peculiar CxxC metal-binding motif, is a protein involved in copper trafficking to the Golgi organelle, allowing the delivery of copper ions to specific target proteins in the secretory pathway. Various experimental studies suggest that this cysteine-containing protein may also be implicated in Zn(II) ion binding, functioning as a regulatory factor. However, despite the relevance of this interaction, the mechanism of zinc ion dissociation from the Atox1 homodimer has not yet been elucidated.We herein devised a simulation workflow based on the combination of parallel bias metadynamics and free energy perturbation theory, to predict the dissociation of the zinc ion from the Atox1 homodimer. The freeenergy simulations successfully predicted the dissociation free energy differences of the process, disclosing a mechanism based on a stepwise dissociation pathway of the zinc ion from the Atox1 homodimer.

Copper homeostasis and the ubiquitin proteasome system.

Copper is involved in many physiological pathways and important biological processes as a cofactor of several copper-dependent enzymes. Given the requirement for copper and its potential toxicity, intracellular copper levels are tightly controlled. Disturbances of human copper homeostasis are characterized by disorders of copper overload (Wilson's disease) or copper deficiency (Menkes disease). The maintenance of cellular copper levels involves numerous copper transporters and copper chaperones. Recently, accumulating evidence has revealed that components of the ubiquitin proteasome system (UPS) participate in the posttranslational regulation of these proteins, suggesting that they might play a role in maintaining copper homeostasis. Cellular copper levels could also affect the activity of the UPS, indicating that copper homeostasis and the UPS are interdependent. Copper homeostasis and the UPS are essential to the integrity of normal brain function and while separate links between neurodegenerative diseases and UPS inhibition/copper dyshomeostasis have been extensively reported, there is growing evidence that these two networks might contribute synergistically to the occurrence of neurodegenerative diseases. Here, we review the role of copper and the UPS in the development of Alzheimer's disease, Parkinson's disease, and amyotrophic lateral sclerosis, and discuss the genetic interactions between copper transporters/chaperones and components of the UPS.

Copper ions inhibit Streptococcus mutans–Veillonella parvula dual biofilm by activating Streptococcus mutans reactive nitrogen species

BackgroundTo investigate the inhibition mechanism of copper ions on Streptococcus mutans–Veillonella parvula dual biofilm.MethodsS. mutans–V. parvula dual biofilm was constructed and copper ions were added at different concentrations. After the biofilm was collected, RNA-seq and qRT-PCR were then performed to get gene information.ResultsThe coculture of S. mutans and V. parvula formed a significantly better dual biofilm of larger biomass than S. mutans mono biofilm. And copper ions showed a more significant inhibitory effect on S. mutans–V. parvula dual biofilm than on S. mutans mono biofilm when copper ions concentration reached 100 µM, and copper ions showed a decreased inhibitory effect on S. gordonii–V. parvula dual biofilm and S. sanguis–V.parvula dual biofilm than on the two mono biofilms as the concentration of copper ions increased. And common trace elements such as iron, magnesium, and zinc showed no inhibitory effect difference on S. mutans–V. parvula dual biofilm. The RNA-seq results showed a significant difference in the expression of a new ABC transporter SMU_651c, SMU_652c, SMU_653c, and S. mutans copper chaperone copYAZ. SMU_651c, SMU_652c, and SMU_653c were predicted to function as nitrite/nitrate transporter-related proteins, which suggested the specific inhibition of copper ions on S. mutans–V. parvula dual biofilm may be caused by the activation of S. mutans reactive nitrogen species.ConclusionsStreptococcus mutans and Veillonella parvula are symbiotic, forming a dual biofilm of larger biomass to better resist the external antibacterial substances, which may increase the virulence of S. mutans. While common trace elements such as iron, magnesium, and zinc showed no specific inhibitory effect on S. mutans–V. parvula dual biofilm, copper ion had a unique inhibitory effect on S. mutans–V. parvula dual biofilm which may be caused by activating S. mutans RNS when copper ions concentration reached 250 µM.

Molecular Interactions of the Copper Chaperone Atx1 of Paracoccidioides brasiliensis with Fungal Proteins Suggest a Crosstalk between Iron and Copper Homeostasis.

Paracoccidioides spp. are endemic fungi from Latin America that cause Paracoccidioidomycosis, a systemic disease. These fungi present systems for high-affinity metal uptake, storage, and mobilization, which counteract host nutritional immunity and mitigate the toxic effects of metals. Regarding Cu mobilization, the metallochaperone Atx1 is regulated according to Cu bioavailability in Paracoccidioides spp., contributing to metal homeostasis. However, additional information in the literature on PbAtx1 is scarce. Therefore, in the present work, we aimed to study the PbAtx1 protein-protein interaction networks. Heterologous expressed PbAtx1 was used in a pull-down assay with Paracoccidioides brasiliensis cytoplasmic extract. Nineteen proteins that interacted with PbAtx1 were identified by HPLC-MSE. Among them, a relevant finding was a Cytochrome b5 (PbCyb5), regulated by Fe bioavailability in Aspergillus fumigatus and highly secreted by P. brasiliensis in Fe deprivation. We validated the interaction between PbAtx1-PbCyb5 through molecular modeling and far-Western analyses. It is known that there is a relationship between Fe homeostasis and Cu homeostasis in organisms. In this sense, would PbAtx1-PbCyb5 interaction be a new metal-sensor system? Would it be supported by the presence/absence of metals? We intend to answer those questions in future works to contribute to the understanding of the strategies employed by Paracoccidioides spp. to overcome host defenses.

Copper trafficking systems in cells: insights into coordination chemistry and toxicity.

Transition metal ions, such as copper, are indispensable components in the biological system. Copper ions which primarily exist in two major oxidation states Cu(I) and Cu(II) play crucial roles in various cellular processes including antioxidant defense, biosynthesis of neurotransmitters, and energy metabolism, owing to their inherent redox activity. The disturbance in copper homeostasis can contribute to the development of copper metabolism disorders, cancer, and neurodegenerative diseases, highlighting the significance of understanding the copper trafficking system in cellular environments. This review aims to offer a comprehensive overview of copper homeostatic machinery, with an emphasis on the coordination chemistry of copper transporters and trafficking proteins. While copper chaperones and the corresponding metalloenzymes are thoroughly discussed, we also explore the potential existence of low-molecular-mass metal complexes within cellular systems. Furthermore, we summarize the toxicity mechanisms originating from copper deficiency or accumulation, which include the dysregulation of oxidative stress, signaling pathways, signal transduction, and amyloidosis. This perspective review delves into the current knowledge regarding the intricate aspects of the copper trafficking system, providing valuable insights into potential treatment strategies from the standpoint of bioinorganic chemistry.

Crossroads between copper ions and amyloid formation in Parkinson's disease.

Copper (Cu) ion dys-homeostasis and α-synclein amyloid deposits are two hallmarks of Parkinson's disease (PD). Here, I will discuss the connections between these features, with a major focus on the role of Cu in the α-synuclein (aS) amyloid formation process. The structurally disordered aS monomer can bind to both redox states of Cu (i.e., oxidized Cu(II) and reduced Cu(I)) with high affinity in vitro. Notably, the presence of Cu(II) (in absence of aS N-terminal acetylation) and Cu(I) (when in complex with the copper chaperone Atox1) modulate aS assembly into β-structured amyloids in opposite directions in vitro. Albeit the link to biological relevance is not fully unraveled, existing observations clearly emphasize the need for more knowledge on this interplay and its consequences to eventually combat destructive reactions that promote PD.

Circ-CCS regulates oxaliplatin resistance via targeting miR-874-3p/HK2 axis in colorectal cancer.

Colorectal cancer (CRC) is a malignancy that threatens the patient's life. Previous reports showed that circular RNAs (circRNAs) can affect CRC development. Herein, we demonstrated the characters of circular RNA copper chaperone for superoxide dismutase (circ-CCS) in CRC tissues and cells. Circ-CCS, CCS mRNA, microRNA-874-3p (miR-874-3p) and hexokinase 2 (HK2) were indicated by qRT-PCR and western blot in CRC. The cell roles were examined. Additionally, the interaction between miR-874-3p and circ-CCS or HK2 was forecasted by the bioinformatics method and assessed by dual-luciferase reporter assay. Finally, the mouse test was implemented to demonstrate the effect of circ-CCS in vivo. Circ-CCS and HK2 were increased, whereas miR-874-3p was diminished in CRC. Circ-CCS lack subdued the IC₅₀ value of oxaliplatin, cell proliferation, migration, invasion and glycolysis metabolism in CRC cells, while it endorsed cell apoptosis. Furthermore, miR-874-3p was validated as having a tumor repressive effect in CRC cells by restraining HK2. The results also showed that HK2 could regulate the development of CRC. In mechanism, circ-CCS targeted miR-874-3p to control HK2. In addition, circ-CCS knock-down also attenuated tumor growth in mice. Circ-CCS expedited CRC through miR-874-3p/HK2.

Zinc-mediated interaction of copper chaperones through their heavy-metal associated domains

BackgroundA copper chaperone CCS is a multi-domain protein that supplies a copper ion to Cu/Zn-superoxide dismutase (SOD1). Among the domains of CCS, the N-terminal domain (CCSdI) belongs to a heavy metal-associated (HMA) domain, in which a Cys-x-x-Cys (CxxC) motif binds a heavy metal ion. It has hence been expected that the HMA domain in CCS has a role in the metal trafficking; however, the CxxC motif in the domain is dispensable for supplying a copper ion to SOD1, leaving an open question on roles of CCSdI in CCS. MethodsTo evaluate protein-protein interactions of CCS through CCSdI, yeast two-hybrid assay, a pull-down assay using recombinant proteins, and the analysis with fluorescence resonance energy transfer were performed. ResultsWe found that CCS specifically interacted with another copper chaperone HAH1, a HMA domain protein, through CCSdI. The interaction between CCSdI and HAH1 was not involved in the copper supply from CCS to SOD1 but was mediated by a zinc ion ligated with Cys residues of the CxxC motifs in CCSdI and HAH1. ConclusionWhile physiological significance of the interaction between copper chaperones awaits further investigation, we propose that CCSdI would have a role in the metal-mediated interaction with other proteins including heterologous copper chaperones.

Molecular manipulations of miR398 increase rice grain yield under different conditions.

Rice miR398 targets two stress-tolerant genes, CSD1-2 (Cu/Zn Superoxide Dismutases1-2) and CCS (copper chaperone of CSD), which usually boost plants' tolerance by inhibiting growth. So, how to accurately regulate the activities of miR398 targets and thus make rice better able to adapt to different conditions has great significances in producing rice yields under the current circumstances of shrinking arable lands resulting from global urbanization and increasing salty soil caused by irrigation. Through controlling the expressions of miR398 in different levels, we found down-regulated expression of miR398 targets can promote growth under good growth conditions while up-regulated expressions of the targets can help rice tolerate salt. In this study, we over-expressed miR398 highly, moderately, and lowly, then three concomitantly inverse levels of its targets' expression were obtained. Under normal growth conditions, the transgenic lines with low and moderate levels of over-expressions of miR398 could increase grain yields 14.5% and 7.3%, respectively, although no transgenic lines could survive well under salty conditions simulating real saline-alkali soil. Using short tandem target mimic (STTM) technology to silence miR398 highly, moderately, and lowly respectively, also three inverse levels of its targets' expression were obtained. All three transgenic lines exhibited good agronomic performances under salt stress in inverse to their degrees of STTM, but their growth was inhibited differently under normal conditions. Altogether, we suggest that flexibly manipulating the expression of miR398 is an ideal strategy to help rice survive better and achieve optimized yields under specific conditions.

H3K27 acetylation activated-CCS regulates autophagy and apoptosis of lung cancer by alleviating oxidative stress

Copper chaperone for superoxide dismutase (CCS) is abnormally expressed in various human malignancies. However, the function and mechanism of CCS in lung cancer progression remain unclear. In the current study, CCS protein and mRNA levels were found to be increased in lung adenocarcinoma (LUAD) tissue and cell lines. Patients with higher CCS levels had a poorer prognosis. Decreasing the enrichment of histone H3 Lys27 acetylation (H3K27ac) by A-485 inhibited CCS expression. CCS depletion upregulated reactive oxygen species (ROS) levels, aggravated oxidative stress, inhibited autophagy, inhibited cell survival, and promoted apoptosis. The treatment of antioxidant N-Acetyl-L-cysteine (NAC) rescued these changes induced by CCS depletion. CCS also was found to be related to the immune infiltration of CD8 + T cells and regulatory T cells in LUAD. These data indicated that overexpression of CCS activated by H3K27 acetylation relieved oxidative stress, promoted autophagy, and inhibited apoptosis. CCS may be regarded as a potential therapeutic target for LUAD.

Integrated Multi-Omic Characterization of the Detachment Process of Adherent Vero Cells with Animal-Based and Animal-Origin-Free Enzymes.

Cell detachment techniques using animal-derived enzymes are necessary for the production of biopharmaceuticals that are made with the help of adherent cell cultures, although the majority of protein therapeutics (>USD 100 billion of income per year) are made under suspension cultures that do not require animal-derived proteins for manufacture. In this study, we establish the optimal Vero cell detachment process, and analyze physiological changes during cell detachment at the cellular and molecular levels. Using flow cytometry, we find that animal-based enzymes are more likely to induce apoptosis than animal-origin-free enzymes. We analyze the levels of RNAs, proteins, and metabolites in cells treated with two detachment strategies, and identify 1237 differentially expressed genes, 2883 differential proteins, and 210 differential metabolites. Transcriptomic analysis shows that animal-origin-free enzymes have a less significant effect on gene expression levels. Combined with proteomic analysis, animal-based enzymes affect the oxidative phosphorylation process and reduce the mRNA and protein levels of Cytochrome C Oxidase Assembly Protein 17 (COX17), which is a Cytochrome C Oxidase Copper Chaperone involved in the mitochondrial respiratory chain. Metabolomics analysis indicates that the levels of spermine and spermidine, which are involved in the glutathione metabolism pathway and apoptosis inhibition, are significantly reduced. Therefore, COX17, spermine, and spermidine may be biomarkers for evaluating the cell subculture process. In conclusion, we have deeply characterized the cell subculture process through multi-omics, which may provide important guidance for research and process evaluation to optimize cell detachment processes.

Crystal Structure of the Human Copper Chaperone ATOX1 Bound to Zinc Ion.

The bioavailability of copper (Cu) in human cells may depend on a complex interplay with zinc (Zn) ions. We investigated the ability of the Zn ion to target the human Cu-chaperone Atox1, a small cytosolic protein capable of anchoring Cu(I), by a conserved surface-exposed Cys-X-X-Cys (CXXC) motif, and deliver it to Cu-transporting ATPases in the trans-Golgi network. The crystal structure of Atox1 loaded with Zn displays the metal ion bridging the CXXC motifs of two Atox1 molecules in a homodimer. The identity and location of the Zn ion were confirmed through the anomalous scattering of the metal by collecting X-ray diffraction data near the Zn K-edge. Furthermore, soaking experiments of the Zn-loaded Atox1 crystals with a strong chelating agent, such as EDTA, caused only limited removal of the metal ion from the tetrahedral coordination cage, suggesting a potential role of Atox1 in Zn metabolism and, more generally, that Cu and Zn transport mechanisms could be interlocked in human cells.

Intestinal mucin is a chaperone of multivalent copper

Mucus protects the epithelial cells of the digestive and respiratory tracts from pathogens and other hazards. Progress in determining the molecular mechanisms of mucus barrier function has been limited by the lack of high-resolution structural information on mucins, the giant, secreted, gel-forming glycoproteins that are the major constituents of mucus. Here, we report how mucin structures we determined enabled the discovery of an unanticipated protective role of mucus: managing the toxic transition metal copper. Using two juxtaposed copper binding sites, one for Cu2+ and the other for Cu1+, the intestinal mucin, MUC2, prevents copper toxicity by blocking futile redox cycling and the squandering of dietary antioxidants, while nevertheless permitting uptake of this important trace metal into cells. These findings emphasize the value of molecular structure in advancing mucosal biology, while introducing mucins, produced in massive quantities to guard extensive mucosal surfaces, as extracellular copper chaperones.