Combining
Ultrarapid Mixing with Photochemical Oxidation
to Probe Protein Folding
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Abstract
We demonstrate a
new method to study protein folding by combining
fast photochemical oxidation of proteins (FPOP) with ultrarapid microfluidic
mixing to observe kinetics on the microsecond time scale. Folding
proteins pass through a focused UV laser beam, creating OH radicals
that label the select protein side chains and are analyzed with mass
spectrometry. As a proof of principle, we demonstrate this method
with hen egg lysozyme that shows at least two kinetic phases before
1 ms, which are compared with those observed by Trp fluorescence.
This method provides another, complementary probe of the early, complex
steps of protein folding