PubMed Health⌕ Search

Biomedical subjects

M D Harrison

Publications and source records attributed to M D Harrison.

11 recordsLinked to original sources
PubMed2000

Stoichiometry of complex formation between Copper(I) and the N-terminal domain of the Menkes protein.

The inherent cellular toxicity of copper ions demands that their concentration be carefully controlled. The cellular location of the Menkes ATPase, a key element in the control of intracellular copper, is regulated by the intracellular copper concentration through the N-terminus of the enzyme, comprising 6 homologous subdomains or modules, each approximately 70 residues in length and containing a -Cys-X-X-Cys- motif. Based on the proposal that binding of copper to these modules regulates the Menkes ATPase cellular location by promoting changes in the tertiary structure of the enzyme, we have expressed the entire N-terminal domain (MNKr) and the second metal-binding module (MNKr2) of the Menkes protein in E. coli and purified them to homogeneity. Ultraviolet-visible, luminescence, and X-ray absorption spectroscopy show that copper and silver bind to the single module, MNKr2, with a stoichiometry of one metal ion per module. However, the array of six modules, MNKr, binds Cu(I) to produce a homogeneous conformer with 4 mol equiv of metal ion. The metal ions are bound in an environment that is shielded from solvent molecules. We suggest a model of the Menkes protein in which the Cu(I) binding induces tertiary changes in the organization of the six metal-binding domains.

Adenosine Triphosphatases↗
PubMed2000

Intracellular copper routing: the role of copper chaperones.

Copper is required by all living systems. Cells have a variety of mechanisms to deal with this essential, yet toxic trace element. A recently discovered facet of homeostatic mechanisms is the protein-mediated, intracellular delivery of copper to target proteins. This routing is accomplished by a novel class of proteins, the 'copper chaperones'. They are a family of conserved proteins present in prokaryotes and eukaryotes, which suggests that copper chaperones are used throughout nature for intracellular copper routing.

Animals↗
PubMed1999

Characterisation of copper-binding to the second sub-domain of the Menkes protein ATPase (MNKr2).

The Menkes ATPase (MNK) has an essential role in the translocation of copper across cellular membranes. In a complementary manner, the intracellular concentration of copper regulates the activity and cellular location of the ATPase through its six homologous amino-terminal domains. The roles of the six amino-terminal domains in the activation and cellular trafficking processes are unknown. Understanding the role of these domains relies on the development of an understanding of their metal-binding properties and structural properties. The second conserved sub-domain of MNK was over-expressed, purified and its copper-binding properties characterised. Reconstitution studies demonstrate that copper binds to MNKr2 as Cu(I) with a stoichiometry of one copper per domain. This is the first direct evidence of copper-binding to the MNK amino-terminal repeats. Circular dichroism studies suggest that the binding or loss of copper to MNKr2 does not cause substantial changes to the secondary structure of the protein.

Adenosine Triphosphatases↗
PubMed1999

The Enterococcus hirae copper chaperone CopZ delivers copper(I) to the CopY repressor.

Expression of the cop operon which effects copper homeostasis in Enterococcus hirae is controlled by the copper responsive repressor CopY. Purified Zn(II)CopY binds to a synthetic cop promoter fragment in vitro. Here we show that the 8 kDa protein CopZ acts as a copper chaperone by specifically delivering copper(I) to Zn(II)CopY and releasing CopY from the DNA. As shown by gel filtration and luminescence spectroscopy, two copper(I) are thereby quantitatively transferred from Cu(I)CopZ to Zn(II)CopY, with displacement of the zinc(II) and transfer of copper from a non-luminescent, exposed, binding site in CopZ to a luminescent, solvent shielded, binding site in CopY.

Amino Acid Sequence↗
PubMed1999

Molecular mechanisms of copper metabolism and the role of the Menkes disease protein.

Menkes disease is an X-linked, recessive disorder of copper metabolism that occurs in approximately 1 in 200,000 live births. The condition is characterized by skeletal abnormalities, severe mental retardation, neurologic degeneration, and patient mortality in early childhood. The symptoms of Menkes disease result from a deficiency of serum copper and copper-dependent enzymes. A candidate gene for the disease has been isolated and designated MNK. The MNK gene codes for a P-type cation transporting ATPase, based on homology to known P-type ATPases and in vitro experimentation. cDNA clones of MNK in Menkes patients show diminished or absented hybridization in northern blot experiments. The Menkes protein functions to export excess intracellular copper and activates upon Cu(I) binding to the six metal-binding repeats in the amino-terminal domain. The loss of Menkes protein activity blocks the export of dietary copper from the gastrointestinal tract and causes the copper deficiency associated with Menkes disease. Each of the Menkes protein amino-terminal repeats contains a conserved -X-Met-X-Cys-X-X-Cys- motif (where X is any amino acid). These metal-binding repeats are conserved in other cation exporting ATPases involved in metal metabolism and in proteins involved in cellular defense against heavy metals in both prokaryotes and eukaryotes. An overview of copper metabolism in humans and a discussion of our understanding of the molecular basis of cellular copper homeostasis is presented. This forms the basis for a discussion of Menkes disease and the protein deficit in this disease.

Adenosine Triphosphatases↗
PubMed1999

Copper chaperones: function, structure and copper-binding properties.

Copper is an absolute requirement for living systems and the intracellular trafficking of this metal to copper-dependent proteins is fundamental to normal cellular metabolism. The copper chaperones perform the dual functions of trafficking and the prevention of cytoplasmic exposure to copper ions in transit. Only a small number of copper chaperones have been identified at this time but their conservation across plant, bacterial and animal species suggests that the majority of living systems utilise these proteins for copper routing. The available data suggest that each copper-dependent protein in the cell is served by a specific copper chaperone. Although copper chaperones cannot be substituted for one another in a given cell type, copper chaperones that deliver to the same protein in different cell types appear to be functionally equivalent. The majority of the copper chaperones identified thus far have an "open-faced beta-sandwich" global fold with a conserved MXCXXC metal-binding motif. Specificity for a given copper-dependent protein appears to be mediated by the residues surrounding the copper-binding motif. Copper binds to such proteins as Cu(I) in a trigonal complex with three sulfur ligands. Only the copper chaperone specific for cytochrome-c-oxidase, Cox17, deviates from this design.

Amino Acid Sequence↗
PubMed1998

Mechanisms for protection against copper toxicity.

Essential transition metals such as copper, molybdenum, and zinc and nonessential metals like cadmium, mercury, and lead can be toxic at the cellular, tissue, and organ levels when present in excess. To avoid metal-induced toxicity most organisms use a redundant combination of metal-regulated import inhibition, sequestration, and enhanced export mechanisms. Combinations of these mechanisms are used to form detoxification pathways controlled through metal-binding proteins at transcriptional, translational, or enzymatic levels. In mammalian pathways copper is partially detoxified by sequestration in the metal-binding metallothioneins or export via the copper-translocating ATPases. Copper regulation of these two mechanisms is afforded by specific conformational changes induced in regulatory proteins on metal binding.

Adenosine Triphosphatases↗
PubMed1986

The "urge to classify" the drug user: a review of classifications by pattern of abuse.

Attempts to classify drug abusers are divided into three main categories: psychiatric classifications, psychosocial classifications and classifications by pattern of abuse. The present article focuses on pattern of abuse classifications which are divided into two subcategories: by substance of abuse and by degree of involvement. The various groups of these classifications are reviewed and their potential uses are discussed. The review indicates that classifications by substance of abuse may be useful for administrative and theoretical purposes, while their clinical uses are limited to medical emergencies. On the other hand, classifications by degree of involvement are useful for initial treatment planning and for predicting treatment outcomes. The authors conclude that for a thorough treatment planning and for better understanding of the addiction process, psychosocial classifications may be the most useful approach.

Humans↗