1. Rhodopsin-cyclases for photocontrol of cGMP/cAMP and 2.3 Å structure of the adenylyl cyclase domain.
- Author
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Scheib U, Broser M, Constantin OM, Yang S, Gao S, Mukherjee S, Stehfest K, Nagel G, Gee CE, and Hegemann P
- Subjects
- Adenylyl Cyclases genetics, Adenylyl Cyclases metabolism, Animals, Binding Sites, Blastocladiomycota chemistry, Blastocladiomycota genetics, Crystallization, Cyclic AMP metabolism, Cyclic GMP metabolism, Fungal Proteins metabolism, Guanylate Cyclase genetics, Guanylate Cyclase metabolism, Models, Molecular, Protein Binding, Protein Domains, Rats, Rhodopsin metabolism, Adenylyl Cyclases chemistry, Blastocladiomycota enzymology, Cyclic AMP chemistry, Cyclic GMP chemistry, Fungal Proteins chemistry, Guanylate Cyclase chemistry, Rhodopsin chemistry
- Abstract
The cyclic nucleotides cAMP and cGMP are important second messengers that orchestrate fundamental cellular responses. Here, we present the characterization of the rhodopsin-guanylyl cyclase from Catenaria anguillulae (CaRhGC), which produces cGMP in response to green light with a light to dark activity ratio >1000. After light excitation the putative signaling state forms with τ = 31 ms and decays with τ = 570 ms. Mutations (up to 6) within the nucleotide binding site generate rhodopsin-adenylyl cyclases (CaRhACs) of which the double mutated YFP-CaRhAC (E497K/C566D) is the most suitable for rapid cAMP production in neurons. Furthermore, the crystal structure of the ligand-bound AC domain (2.25 Å) reveals detailed information about the nucleotide binding mode within this recently discovered class of enzyme rhodopsin. Both YFP-CaRhGC and YFP-CaRhAC are favorable optogenetic tools for non-invasive, cell-selective, and spatio-temporally precise modulation of cAMP/cGMP with light.
- Published
- 2018
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