Kinetic analysis of the interaction of the copper chaperone Atox1 with the metal binding sites of the Menkes protein

Excess copper is effluxed from mammalian cells by the Menkes or Wilson P-type ATPases (MNK and WND, respectively). MNK and WND have six metal binding sites (MBSs) containing a CXXC motif within their N-terminal cytoplasmic region. Evidence suggests that copper is delivered to the ATPases by Atox1, one of three cytoplasmic copper chaperones. Attempts to monitor a direct Atox1-MNK interaction and to determine kinetic parameters have not been successful. Here we investigated interactions of Atox1 with wild-type and mutated pairs of the MBSs of MNK using two different methods: yeast two-hybrid analysis and real-time surface plasmon resonance (SPR). A copper-dependent interaction of Atox1 with the MBSs of MNK was observed by both approaches. Cys to Ser mutations of conserved CXXC motifs affected the binding of Atox1 underlining the essentiality of Cys residues for the copper-induced interaction. Although the yeast two-hybrid assay failed to show an interaction of Atox1 with MBS5/6, SPR analysis clearly demonstrated a copper-dependent binding with all six MBSs highlighting the power and sensitivity of SPR as compared with other, more indirect methods like the yeast two-hybrid system. Binding constants for copper-dependent chaperone-MBS interactions were determined to be 10–5-10–6 M for all the MBSs representing relatively low affinity binding events. The interaction of Atox1 with pairs of the MBSs was non-cooperative. Therefore, a functional difference of the MBSs in the MNK N terminus cannot be attributed to cooperativity effects or varying affinities of the copper chaperone Atox1 with the MBSs.

Feudalism in the service of the revolution Reclaiming heritage in hue

This article discusses the way the past is being reexamined in modern-day Vietnam, particularly through the medium of heritage. Hue, the old royal capital of Vietnam, provides the case study, as this city reflects the great themes and events of Vietnamese history over the last two hundred years, from the establishment of a unified nation under the Nguyen, through the imposition of colonial control, the devastation of war, reunification, and the establishment of communism, to the consolidation of an independent postcolonial nation. The importance of Hue's heritage is recognized in its status as a “world heritage” site. The author argues that Hue's heritage is, nevertheless, problematic for Vietnam's ruling communists, because to them it largely represents a regime—the Nguyen Dynasty—that was “reactionary” and that had sold out the country to the French. The apparent contradiction between the standard communist view of the Nguyen past and the value accorded to Nguyen heritage in Hue is resolved, the author contends, by recourse to the depoliticized practices of heritage preservation and tourist promotion.

Are the earliest field monuments of the pacific landscape serial sites?

Explanations of the origin and genesis of Pacific field monuments commonly assume they reflect local social change in islands or island groups which were increasingly isolated following colonization. A recent review of early West Polynesian archaeology suggests that the penecontemporaneous appearance of various kinds of field monuments from eastern Melanesia to Polynesia may be better explained as evidence of interaction and the movement of people and/or ideas, possibly associated with the colonization of East Polynesia.

Learning through computer-mediated communication: a comparison of Australian and Chinese heritage students

Twelve Australian and 12 Chinese heritage students from a third-year university computer ethics subject completed a Readiness for Online Learning Questionnaire; and six students from each of these two groups participated in a student-facilitated problem-solving discussion through computer-mediated communication. The questionnaire comparisons showed that the two groups of students were equally willing to self-manage their own learning, but that Australian students were significantly more comfortable with e-learning. The analysis of student postings in the CMC component showed that, collectively, Australian students posted more messages than did the Chinese students. Both groups participated equally in socialisation online; although Chinese heritage students posted a higher number of messages associated with organisational matters; and Australian students posted a larger number of message components associated with intellectual contributions to the discussion. These results are interpreted in a theoretical context and implications for practice are drawn.