Many organisms rely on a superfamily of flavoproteins to sense light as photoreceptors for signal transduction such as cryptochrome and as photoenzymes for DNA repair such as photolyases, yet it is largely unknown how these proteins can perform their dual functions. We developed a label-free single-molecule detection method based on graphene-molecule-graphene single-molecule junctions to electrically "listen" to the response of a single CraCRY molecule, a bifunctional cryptochrome from Chlamydomonas reinhardtii. We detected robust two-level reversible conformational switching driven only by the C-terminal tail; one is long-lived in several milliseconds, and the other is in submilliseconds with more ordered structure. Only the long, stable structure recognizes the damaged DNA substrate with a binding lifetime of tens of milliseconds. With and without blue-light illumination, our results support that only the reduced state (FADH-) is the active state for both signaling and repair functions. These findings directly link redox chemistry, conformational dynamics, and DNA repair and can provide a reliable solution for how a protein performs dual functions under blue-light illumination.
Zhang et al. (Wed,) studied this question.
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