Table 2.
Phenotype | Template | Modification and Mutations | Effect or Function | Ref |
---|---|---|---|---|
Fluorescent proteins and chromoproteins | ||||
Short NIR | PAS-GAF or PAS- GAF-PHY domains | Truncation of PHY domain; Truncation of up to two amino acids before Cys12; 194A,H,K,L,S; 247A | Stabilization of the chromophore in the Pr state with disabling of Pr→Pfr photoconvertion | 32, 38, 41 |
194A,H,K,L,S; 250F; 277Q | Increase in quantum yield | 12, 41, 42 | ||
163H, 185L, 195D, 459A, 453A, 277A,Q | Stabilization of the Pr state with limited/reduced photoconversion | 32, 33, 38, 41 | ||
Long NIR | PAS-GAF-PHY domains of bathy BphPs | 261A | Stabilization of the Pfr state with disabling Pfr→Pr photoconvertion | 38 |
163A; 241A; 275A | Stabilization of the Pfr state with reducing Pfr→Pr photoconvertion | 38 | ||
PS and PA NIR (switching on) | PAS-GAF-PHY domains | 188L; 275A; 190A; 163H; 250F | Decreasing rate of Pr→Pfr dark reversion (from minutes to hours) | 33, 38 |
241A; 163A | Increasing rate of Pr →Pfr dark reversion (faster than 3 min) | 38 | ||
PA NIR (switching off) | PAS-GAF domains | 194A,T,Q; 260A,S | Reversible bleaching of Pr state with no photoconversion to Pfr state | 41 |
Monomeric | PAS-GAF or PAS- GAF-PHY domains | 131S; 295E; 298D,K; 301D,R; 305R | Disruption of the dimer interface | 11, 12 |
Biosensors | ||||
Redox sensor | Optimized BphP- derived FPs | Residues located in close proximity to the thioether linkage between BV and apoprotein | Catalyzing thioether bond formation and influencing its reactivity | 41, 58 |
Metal sensor | PAS-GAF domains | Truncation of PHY domain | Increasing solvent access to chromophore | 13, 32, 34, 59 |
Residues within 4.5Å from the chromophore | Improving interactions between metal ion and chromophore | |||
Split and insertion based sensors * | Optimized BphP- derived FPs | Split/insertion between 112–119 amino acid residues | Unstructured linker between PAS and GAF domains | 32–34, 38, 60, 61 |
Varying the linkers between PAS domain and sensing moiety, and GAF and sensing moiety | Optimization of PAS and GAF domains collocation for their better interactions | |||
Optogenetic tools | ||||
Optogenetic tools with different effector modules | PAS-GAF-PHY domains of BphP and a knowledge-based chosen effector module | Varying the α-helix linker between photosensor and effector modules | Ability of light signal propagation to effector | 18, 62, 63 |
Point mutations in the α-helix linker and PAS domain | Efficiency of light signal propagation to effector | 64 | ||
188L; 275A; 190A; 163H; 250F; 241A; 163A | Optimization of photopreception | 32, 33, 38, 47 |
Structure of the PAS-GAF domains contains a 4-crossover knot that may complicate reconstitution of a split protein. Residues at the indicated positions provide the respective phenotype in concerted manner or independently. Residue numbering follows that for PaBphP. See Fig. 3 for the amino acid alignment of several BphPs.