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Team:Tokyo-NoKoGen/rhodopsin
From 2013.igem.org
Improving a BioBrick part
Sensory Rhodopsin fused with EnvZ histidine kinase (BBa_K769000)
From Tokyo-NoKoGen 2012 https://2012.igem.org/Team:Tokyo-NoKoGen
Fig.1 Sensory rhodopsin II (SRII) (orange) from N. pharaonis, fused to cognate transducer protein HtrII (green) by a 9 amino acid linker, fused to E. coli chemotaxis receptor Tsr (blue) in the cytoplasmic region. (Ref.4) Fig.2 Construct from Tokyo-NoKogen 2012. EnvZ hisidine kinase domain was obtained from cph8, cut at NdeI and SpeI and fused together.
Last year's team member ofTokyo-NoKoGen2012 have been working on developing a light sensor by combining the sensory domain of rhodospin derived from N. pharaonis, and the histidine kinase domain of EnvZ from Escherichia coli.
Halophilic archaea, such as Halobacterium salinarum and Natronobacterium pharaonis (N. pharaonis) are known to show phototaxis by the illumination of light by using receptors called sensory rhodopsin I and II (SRI and SRII). The SR proteins are seven-transmembrane retinylidene photoreceptors, which transmits blue light signal (λmax 487 nm) to their corresponding transducers HtrI and HtrII respectively. This two-component system regulates cells’ flagellar motors for phototaxis (Ref. 1, 2).
It has been previously reported that chimeric rhodopsin can be constructed inside E. coli. Kwang-Hwan et al. constructed chimeric transducer proteins, by exchanging the Histidine-kinase domain of the bacteriorhodopsin from N. pharaonis with homologous chemotaxis transducers Tsr and Tar of E. coli and Salmonella enterica, (chemotaxis transducers of serine and aspartate respectively).They observed a that E. coli succesfully responded to light (Ref.3). It was also previously reported, that the sensory domain of Tar protein fused with histidine kinase domain of EnvZ protein of E. coli, functioned as a sensor protein to transmit signal received by the Tar sensory domain to the EnvZ two-component system (Ref.5). The histidine kinase domain of Tar protein and EnvZ protein are homologous.
So last year, Tokyo-NoKoGen 2012 have deduced that sensory domain of sensory rhodopsin fused with EnvZ protein should also function to transmit signal downstream of the two-component system.
Reference
[1] Xue-Nong Zhang et al. (1999) The specificity of interaction of archaeal transducers with their cognate sensory rhodopsins is determined by their transmembrane helices, Proc. Natl. Acad. Sci. USA
[2] Wouter D. Hoff et al. (1997) Molecular mechanism of photosignaling by archaeal sensory rhodopsin, Anmu. Rev. Biophys. Biomol. Struct.
[3] Kwang-Hwan Jung et al. (2001) An archaeal photosignal-transducing module mediates phototaxis in Escherichia coli, Journal of bacteriology
[4] Vishwa D. et al. (2003) Photostimulation of a sensory rhodopsin II/HtrII/Tsr fusion chimera activates CheA-autophosphorylation and CheY-phototransfer in vitro, Biochemistry
[5] Yoshida T et al., (2007) The design and development of Tar-EnvZ chimeric receptors, Methods Enzymol.
