Finding folding flaws
Understanding the mechanisms behind protein folding is one of the remaining challenges for protein chemists, and this challenge becomes particularly important when you consider that human diseases like Alzheimer’s and Creutzfeld-Jakob disease are largely
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LEEDS, U.K.—Understanding the mechanisms behind protein folding is one of the remaining challenges for protein chemists, and this challenge becomes particularly important when you consider that human diseases like Alzheimer's and Creutzfeld-Jakob disease are largely the result of misfolded proteins or peptides. Recently, researchers at the University of Leeds and the Waters MS Technologies Centre examined folding using the complementary MS methods: electrospray ionization (ESI-) MS and ion mobility spectrometry (IMS).
As they reported in the Journal of the ASMS, combined ESI-MS's ability to analyze the mass and ionization state of peptides with IMS's ability to separate ions not just on the basis of m/z but also on the basis of their physical cross-section (or conformation). They used the methods to examine cytochrome c and the amyloidogenic protein β2-microglobulin (β2m). They found that under different buffer conditions, they could identify a variety of conformational states for each protein, representing fully and partially folded as well as denatured protein.
The then examined β2m mutants and noted that although the I7A mutant rapidly denatures, it largely retains a native conformation, whereas the I7A/P32G double mutant has a partially folded conformation, which may explain why the double mutant more rapidly forms amyloid fibrils in vivo. Thus, the researchers have linked molecular conformation with biological behavior.
"We can now quantify the amount of protein that is in its native state and the amount that is unfolded and partially folded," says lead author Dr. Alison Ashcroft. "We can also monitor which particular conformers are consumed during the assembly process. This is providing important new insights and detail into how biomolecules work at the molecular level."
