Borysik, A.J.H., Radford, S.E. and Ashcroft, A.E. (2004) Co-populated Conformational Ensembles of β(2)-Microglobulin Uncovered Quantitatively by Electrospray Ionization Mass Spectrometry. Journal of Biological Chemistry, 279 (26). pp. 27069-27077. ISSN 1083-351X
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Ordered assembly of monomeric human β(2)-microglobulin (β(2)m) into amyloid fibrils is associated with the disorder hemodialysis-related amyloidosis. Previously, we have shown that under acidic conditions (pH <5.0 at 37 °C), wild-type β(2)m assembles spontaneously into fibrils with different morphologies. Under these conditions, β(2)m populates a number of different conformational states in vitro. However, this equilibrium mixture of conformationally different species is difficult to resolve using ensemble techniques such as nuclear magnetic resonance or circular dichroism. Here we use electrospray ionization mass spectrometry to resolve different species of β(2)m populated between pH 6.0 and 2.0. We show that by linear deconvolution of the charge state distributions, the extent to which each conformational ensemble is populated throughout the pH range can be determined and quantified. Thus, at pH 3.6, conditions under which short fibrils are produced, the conformational ensemble is dominated by a charge state distribution centered on the 9+ ions. By contrast, under more acidic conditions (pH 2.6), where long straight fibrils are formed, the charge state distribution is dominated by the 10+ and 11+ ions. The data are reinforced by investigations on two variants of β(2)m (V9A and F30A) that have reduced stability to pH denaturation and show changes in the pH dependence of the charge state distribution that correlate with the decrease in stability measured by tryptophan fluorescence. The data highlight the potential of electrospray ionization mass spectrometry to resolve and quantify complex mixtures of different conformational species, one or more of which may be important in the formation of amyloid.
|Copyright, Publisher and Additional Information:||Copyright © 2004 by the American Society for Biochemistry and Molecular Biology.|
|Institution:||The University of Leeds|
|Academic Units:||The University of Leeds > University of Leeds Research Centres and Institutes > Astbury Centre for Structural Molecular Biology (Leeds)
The University of Leeds > Faculty of Biological Sciences (Leeds) > School of Molecular and Cellular Biology (Leeds)
|Depositing User:||Repository Officer|
|Date Deposited:||13 Mar 2006|
|Last Modified:||18 Aug 2015 05:34|