Institut for Biomedicin

Mikael Esmann

Spin-echo EPR of Na,K-ATPase unfolding by urea

Publikation: Bidrag til tidsskrift/Konferencebidrag i tidsskrift /Bidrag til avisTidsskriftartikelForskningpeer review

  • Rita Guzzi
  • ,
  • Mohammad Babavali
  • ,
  • Rosa Bartucci
  • ,
  • Luigi Sportelli
  • ,
  • Mikael Esmann
  • Derek Marsh
  • Institut for Fysiologi og Biofysik
Denaturant-perturbation and pulsed EPR spectroscopy are combined to probe the folding of the membrane-bound Na,K-ATPase active transport system. The Na,K-ATPase enzymes from shark salt gland and pig kidney are covalently spin labelled on cysteine residues that either do not perturb or are essential to hydrolytic activity (Class I and Class II -SH groups, respectively). Urea increases the accessibility of water to the spin-labelled groups and increases their mutual separations, as recorded by D(2)O interactions from ESEEM spectroscopy and instantaneous spin diffusion from echo-detected EPR spectra, respectively. The greater effects of urea are experienced by Class I groups, which indicates preferential unfolding of the extramembrane domains. Conformational heterogeneity induced by urea causes dispersion in spin-echo phase-memory times to persist to higher temperatures. Analysis of lineshapes from partially relaxed echo-detected EPR spectra indicates that perturbation by urea enhances the amplitude and rate of fluctuations between conformational substates, in the higher temperature regime, and also depresses the glasslike transition in the protein. These non-native substates that are promoted by urea lie off the enzymatic pathway and contribute to the loss of function.
OriginalsprogEngelsk
TidsskriftBBA General Subjects
Vol/bind1808
Sider (fra-til)1618-1628
Antal sider11
ISSN0304-4165
DOI
StatusUdgivet - 1 apr. 2011

Se relationer på Aarhus Universitet Citationsformater

ID: 35256234