Interplay between chromophore binding and domain assembly by the B12-dependent photoreceptor protein, CarH

Organisms across the natural world respond to their environment through the action of photoreceptor proteins. The vitamin B-12-dependent photoreceptor, CarH, is a bacterial transcriptional regulator that controls the biosynthesis of carotenoids to protect against photo-oxidative stress. The binding of B-12 to CarH monomers in the dark results in the formation of a homo-tetramer that complexes with DNA; B-12 photochemistry results in tetramer dissociation, releasing DNA for transcription. Although the details of the response of CarH to light are beginning to emerge, the biophysical mechanism of B-12-binding in the dark and how this drives domain assembly is poorly understood. Here - using a combination of molecular dynamics simulations, native ion mobility mass spectrometry and time-resolved spectroscopy - we reveal a complex picture that varies depending on the availability of B-12. When B-12 is in excess, its binding drives structural changes in CarH monomers that result in the formation of head-to-tail dimers. The structural changes that accompany these steps mean that they are rate-limiting. The dimers then rapidly combine to form tetramers. Strikingly, when B-12 is scarcer, as is likely in nature, tetramers with native-like structures can form without a B-12 complement to each monomer, with only one apparently required per head-to-tail dimer. We thus show how a bulky chromophore such as B-12 shapes protein/protein interactions and in turn function, and how a protein can adapt to a sub-optimal availability of resources. This nuanced picture should help guide the engineering of B-12-dependent photoreceptors as light-activated tools for biomedical applications.

Automated summary

A study using molecular dynamics simulations, native ion mobility mass spectrometry and time-resolved spectroscopy has revealed the mechanism of B-12-dependent photoreceptor CarH in the dark and how B-12 drives domain assembly in CarH. When B-12 is in excess, it can form head-to-tail dimers that quickly combine to form tetramers; while when B-12 is scarce, tetramers can still form without a complete B-12 complement to each dimer.