The photochemical mechanism of a B12-dependent photoreceptor protein

Abstract

The coenzyme B12-dependent photoreceptor protein, CarH, is a bacterial transcriptional regulator that controls the biosynthesis of carotenoids in response to light. On binding of coenzyme B12 the monomeric apoprotein forms tetramers in the dark, which bind operator DNA thus blocking transcription. Under illumination the CarH tetramer dissociates, weakening its affinity for DNA and allowing transcription. The mechanism by which this occurs is unknown. Here we describe the photochemistry in CarH that ultimately triggers tetramer dissociation; it proceeds via a cob(III)alamin intermediate, which then forms a stable adduct with the protein. This pathway is without precedent and our data suggest it is independent of the radical chemistry common to both coenzyme B12 enzymology and its known photochemistry. It provides a mechanistic foundation for the emerging field of B12 photobiology and will serve to inform the development of a new class of optogenetic tool for the control of gene expression.

Insights into the lifestyle of uncultured bacterial natural product factories associated with marine sponges

“…For Ca. Entotheonella, a possible role of the carotenoid could be UV-induced stress defense, because the biosynthesis cluster is flanked by a B 12 -dependent light-sensing transcription factor (40). Thus, Ca.…”

New light on vitamin B12: The adenosylcobalamin-dependent photoreceptor protein CarH

“…Kutta et al 56 propose a variant of Path 2 in which heterolysis of the Co-C bond gives the 5'-deoxyadenosyl anion and either hydridocobalamin or a five-coordinate positively charged Co(III) as intermediates on the route to 4',5'-anhydroadenosine and cob(II)alamin. It would seem that Path 2 would be highly unlikely.…”

“…In an attempt to determine a more detailed photochemical mechanism for CarH, Kutta et al 56 performed photoexcitation experiments on the AdoCbl-bound CarH tetramer (ground-state CarH) and used transient UV-visible absorption spectroscopy on a femtosecond (10 -15 s) to second timescale to follow the intermediates on the way to the light-state CarH product. They observed, in addition to Co(II) (and the presumed 5'-deoxyadenosyl radical), transient absorption spectra of at least eight intermediates and assigned these to possible structures, based on a model of the reaction pathway from ground-state CarH to light-state CarH.…”

“…In their minor pathway, A converts to C, which has a spectrum that can be interpreted as a metal-to-ligand charge transfer complex, consisting of Co(III) with a partial positive charge and an incipient 5'-deoxyadenosyl anion. C then converts to D* (in the text) or C* (in their Figure 6), 56 which is interpreted as a five-coordinate, positively charged Co(III) together with a 5'-deoxyadenosyl anion, then to D, in which the 5'-deoxyadenosyl anion has moved away from Co and has been replaced by His132, and lastly to E, which is similar to light-state CarH. 56 Alternatively, they propose that Intermediate C corresponds to hydridocobalamin.…”

B 12 -dependent photoresponsive protein hydrogels for controlled stem cell/protein release

“…The early work led by Murphy and coworkers (20,21) showed the success of synthesizing calmodulin-based protein hydrogels with dynamic properties responsive to Ca 2+ and the small-molecule ligand trifluoperazine. Recently, the CarH protein, a transcriptional regulator controlling bacterial carotenoid synthesis, has been shown to sense and respond to visible light through its C-terminal adenosylcobalamin binding domain (CarH C ) (22)(23)(24)(25). The CarH C domains tetramerize when binding to adenosylcobalamin (AdoB 12 ) in the dark and can readily dissociate into monomers accompanied with a drastic protein conformational change caused by the cleavage of the C-Co bond on exposure to green (522 nm) or white light (Fig.…”