Structural basis of a light-activated signalling pathway involved in Brucella virulence

JIMENA RINALDI; GASTÓN PARIS; SEBASTIÁN KLINKE; FERNANDO A. GOLDBAUM

Salta

Congreso; 3rd Latin American Protein Society Meeting; 2010

Latin American Protein Society

The photosensor LOV-HK is involved in the virulence process of Brucella abortus mediated by blue light. This protein contains an N-terminal photosensory LOV domain, a PAS domain and a C-terminal histidine kinase. The LOV domains were first described in plant phototropins. They consist of an α/β core where the FMN is bound and a C-terminal α-helix, called the J-helix, which is involved in the transduction of the signal to the output domain of the molecule. In phototropins, light absorption causes formation of a covalent bond between a conserved cysteine and atom C(4a) from the FMN ring. This covalent bond breaks spontaneously after 20-30 s in dark, completing the photocycle. In Brucella LOV-HK the adduct state is extremely stable and does not decay significantly in 2 hours. However, a Brucella LOV domain fragment comprising only the α/β core completes the photocycle in 8 hours. The addition of only 20 amino acids corresponding to the J-helix to this core abrogates the dark recovery, showing the same truncated photocycle as the full-length LOV-HK protein. To understand the structural basis of the photochemical behaviour of Brucella LOV-HK, we determined the crystal structure of a stable fragment comprising the α/β core of the LOV domain. This structure shows the canonical fold previously observed for other LOV domains, comprising a five-stranded antiparallel β-sheet and four α-helices. The comparison of this structure with that of LOV domain structures from other photosensory proteins and the kinetics of the photocycle of the different fragments suggest that the J-helix is not just a transducer of the signal but also it determines the photochemical behavior of the molecule.