Oral 23rd International Society of Magnetic Resonance Conference 2023

Mechanism of Co-C Bond Cleavage in B12-Dependent Photoreceptor and the Intermediates probed by Crystallography, UV-Vis and cw-EPR/annealing Methods (#114)

Muralidharan Shanmugam 1 , Harshwardhan Poddar 1 , Derren J Heyes 1 , David Leys 1 , Nigel S Scrutton 1
  1. The University of Manchester, Manchester, ENGLAND, United Kingdom

Coenzyme B12 binding proteins have gained a lot of attention recently for their ability to respond to light and act as photoreceptors.1 CarH is a bacterial coenzyme B12-dependent photoreceptor involved in regulating the expression of carotenoid biosynthetic genes.1 In the dark, CarH is a tetramer and suppresses transcription by binding to the promoter region. Upon illumination of CarH, the photosensitive Co-C bond in B12 is cleaved, triggering a series of intermediate states which culminates in the light-adapted, monomer formation and release of DNA. The nature of the Co-C bond cleavage (homolytic vs heterolytic) and the different intermediates formed during the photocycle remains poorly characterized.2-4

In this presentation the crystal structures of the native and photo-activated CarH from Thermus thermophilus, TtCBD enzyme and the associated dynamics of the B12-chromophore during the photocatalysis will be discussed.5 The cryo-trapping experiment along with cw-EPR/annealing and spin-trapping techniques revealing the possible intermediates present along the CarH photocycle. The role of the axial H132 ligand in the photocycle and the UV-Vis data of these enzymes will also be discussed in detail.     

 

References:

  1. Badmanabhan, R. Perez-Castano and M. Elias-Arnanz; Curr. Opin. Struct. Biol., 2019, 57,47-55.
  2. J. Kutta, S. J. O. Hardman, L. O. Johannissen, B. Bellina, H. L. Messiha, J. M. Ortiz-Guerrero, M. Elias-Arnanz, S. Padmanabhan, P. Barran, N. S. Scrutton and A. R. Jones; Nat. Commun., 2015, 6:7907.
  3. A. Miller, A. K. Kaneshiro, A. Konar, R. Alonson-Mori, A. Britz, A. Deb, J. M. Glownia, J. D. Koralek, L. Mallik, J. H. Meadows et. al; J. Phys. Chem. B.; 2020, 124, 10732-10738.   
  4. Jost, J. H. Simpson and C. L. Drennan; Biochemistry, 2015, 54, 3232-3234.
  5. Poddar et al., Nature Communications, 2023, under revision.