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Microorganisms convert sunlight into biological signals or energy using microbial rhodopsins via intricate mechanism. Vast family of such microbial rhodopsins are categorized as ion pumps with established model bacteriorhodopsin, proteorhodopsin etc. Well studied sensory rhodopsins from archaea are evident to mediate photo modulated response via interaction with their cognate transducer partner within the transmembrane. Such mechanism is quite distinct from mammalian visual rhodopsins, which primarily involve activation within the membrane and relay signal to cytoplasmic protein. We have proposed a binding model of sensory rhodopsin from Anabaena PCC7120 interacting with cytoplasmic transducer protein. This pattern is analogous to visual rhodopsin, rather than to microbial sensory rhodopsin. Anabaena sensory rhodopsin along with putative transducer is linked to photoinduced chromatic adaptation. The atomic resolution structure of Anabaena sensory rhodopsin 1XIO and its putative transducer 2II7 does not account for much insight towards critical motifs. For instance, the crystal structure of the photosensor excludes the vital domain which is almost 12% of total sequence. The 35 amino acid cytoplasmic domain beyond 226 SRQTTGDRFAENTLQFVENITLFANSRRQQSRRRV missing in crystal structure in primarily helical. It comprises of series of Arg in it. Isothermal titration calorimetry analysis reflected its relatively weaker interaction with affinity of 3.61 x104 stoichiometry of 1:4 further decreased to over 5 fold without cytoplasmic domain. Interestingly the thermodynamic parameters of receptor-transducer participation of electrostatic interaction. The involvement of light-dark adaptation in the presence and absence of transducer and/or cytoplasmic domain of receptor further supports the critical role of this domain in binding. The transducer in dicistronic operon in PCC7120, is a 125 amino acid soluble protein assembled as relatively stable tetramer. The transducer contains lone Trp9 as central to a hold two sheets of the sandwich together. Fluorescence study strongly reflect microenvironment perturbation in presence of lipids suggesting its proximity to membrane. Often tools for protein structure prediction rely solely on homology modeling, which only works well for proteins that have a high degree of sequence similarity to protein sequences in the Protein Data Bank, PDB. By contrast, NovaFold and NovaFold AI algorithm from DeepMind use two unique algorithms to create highly accurate, full 3D atomic models of proteins that are unattainable through standard modeling methodologies. It utilizes the Iterative Threading ASSEmbly Refinement, I-TASSER protein structure prediction algorithm. We use it to reconstruct the motif. Further, charged analysis and molecular docking analysis revealed highly consensus model involving electrostatic interaction between Arg of receptor and Asp of transducer. One of consensus model revealed the possible involvement of Arg234 of receptor to Asp79 of transducer. We are currently working on dynamics study and anticipate to complement the outcome of modular docking study. It is likely that other docking model would outline the plausible role of other charged residues present in cytoplasmic helical domain of receptor in binding to transducer. We plan to include the biochemical pulldown assay results using GST tagged transducer with full length and truncated receptor. Supported by Bethune Cookman University RISE 2023 support to project work and previous support from NIH-NIGMS SCORE SC3GM113803 ended in 2020.
Trivedi et al. (Fri,) studied this question.