348 research outputs found
Characterization of early graft damage after pancreatic transplantation
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The Costs of Separation: Friction Between Company and Insolvency Law in the Single Market
Calorimetric Investigation of Copper Binding in the N-Terminal Region of the Prion Protein at Low Copper Loading: Evidence for an Entropically Favorable First Binding Event
Although
the Cu<sup>2+</sup>-binding sites of the prion protein have been well
studied when the protein is fully saturated by Cu<sup>2+</sup>, the
Cu<sup>2+</sup>-loading mechanism is just beginning to come into view.
Because the Cu<sup>2+</sup>-binding modes at low and intermediate
Cu<sup>2+</sup> occupancy necessarily represent the highest-affinity
binding modes, these are very likely populated under physiological
conditions, and it is thus essential to characterize them in order
to understand better the biological function of copper–prion
interactions. Besides binding-affinity data, almost no other thermodynamic
parameters (e.g., Δ<i>H</i> and Δ<i>S</i>) have been measured, thus leaving undetermined the enthalpic and
entropic factors that govern the free energy of Cu<sup>2+</sup> binding
to the prion protein. In this study, isothermal titration calorimetry
(ITC) was used to quantify the thermodynamic parameters (<i>K</i>, Δ<i>G</i>, Δ<i>H</i>, and <i>T</i>Δ<i>S</i>) of Cu<sup>2+</sup> binding to
a peptide, PrPÂ(23–28, 57–98), that encompasses the majority
of the residues implicated in Cu<sup>2+</sup> binding by full-length
PrP. Use of the buffer <i>N</i>-(2-acetomido)-aminoethanesulfonic
acid (ACES), which is also a well-characterized Cu<sup>2+</sup> chelator,
allowed for the isolation of the two highest affinity binding events.
Circular dichroism spectroscopy was used to characterize the different
binding modes as a function of added Cu<sup>2+</sup>. The <i>K</i><sub>d</sub> values determined by ITC, 7 and 380 nM, are
well in line with those reported by others. The first binding event
benefits significantly from a positive entropy, whereas the second
binding event is enthalpically driven. The thermodynamic values associated
with Cu<sup>2+</sup> binding by the Aβ peptide, which is implicated
in Alzheimer’s disease, bear striking parallels to those found
here for the prion protein
Early Onset Prion Disease from Octarepeat Expansion Correlates with Copper Binding Properties
Insertional mutations leading to expansion of the octarepeat domain of the prion protein (PrP) are directly linked to prion disease. While normal PrP has four PHGGGWGQ octapeptide segments in its flexible N-terminal domain, expanded forms may have up to nine additional octapeptide inserts. The type of prion disease segregates with the degree of expansion. With up to four extra octarepeats, the average onset age is above 60 years, whereas five to nine extra octarepeats results in an average onset age between 30 and 40 years, a difference of almost three decades. In wild-type PrP, the octarepeat domain takes up copper (Cu2+) and is considered essential for in vivo function. Work from our lab demonstrates that the copper coordination mode depends on the precise ratio of Cu2+ to protein. At low Cu2+ levels, coordination involves histidine side chains from adjacent octarepeats, whereas at high levels each repeat takes up a single copper ion through interactions with the histidine side chain and neighboring backbone amides. Here we use both octarepeat constructs and recombinant PrP to examine how copper coordination modes are influenced by octarepeat expansion. We find that there is little change in affinity or coordination mode populations for octarepeat domains with up to seven segments (three inserts). However, domains with eight or nine total repeats (four or five inserts) become energetically arrested in the multi-histidine coordination mode, as dictated by higher copper uptake capacity and also by increased binding affinity. We next pooled all published cases of human prion disease resulting from octarepeat expansion and find remarkable agreement between the sudden length-dependent change in copper coordination and onset age. Together, these findings suggest that either loss of PrP copper-dependent function or loss of copper-mediated protection against PrP polymerization makes a significant contribution to early onset prion disease
Corporate Governance, Capital Market Regulation and the Challenge of Disembedded Markets
Is the Board Neutrality Rule Trivial? Amnesia About Corporate Law in European Takeover Regulation
Significant benefits in survival by the use of surgery combined with radiotherapy for retroperitoneal soft tissue sarcoma
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