ENGINEERING RED-LIGHT ACTIVATED NUCLEOTIDE CYCLASES

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Engineered photoregulated proteins have the potential to revolutionize biomedical research. In a photoregulated protein, a photon absorbed by a chromophore bound to a photoreceptor protein domain affects activity of an output domain. Infrared light is harmless to mammalian cells, therefore, it can work as a highly specific, and cheap way to regulate protein activities. The spatiotemporal resolution that can be achieved by using photoregulated proteins is unprecedented as a laser beam can be focused not only on an individual cell but on a particular region of the cell. Engineered photoregulated proteins can be broadly used for activation (or inactivation) of proteins of interest in cell cultures, tissues and animal models. Thus far only blue-light photoreceptors have been used for protein engineering. Bacteriophytochromes absorb red/near infrared light, which has much higher tissue penetration capacity than blue light and is currently used in deep-tissue phototherapies. The objective of this application is to provide the proof of principle that a chromophore-binding module of bacteriophytochromes can be used for engineering of red/near infrared light regulated proteins. The goal of this pilot project is to engineer a near infrared light activated adenylate cyclase (cAMP synthase). The critical role of cAMP in controlling glucose and lipid metabolism as well as neuronal activity makes adenylate cyclase a highly desired target. The design of photoregulated enzymes relies heavily on computationally-intensive bioinformatics approaches that involve analysis and modeling of protein structures and dynamics.