RS/tRNA Foundational Publication Support
Zhu, Phillip, Philip R Gafken, Ryan A Mehl, and Richard B Cooley. (2019) 2019. “A Highly Versatile Expression System For The Production Of Multiply Phosphorylated Proteins.”. Acs Chemical Biology 14 (7): 1564-1572. doi:10.1021/acschembio.9b00307.
RS/tRNA Usage Publications
Stuber, Katrin, Tobias Schneider, Jill Werner, Michael Kovermann, Andreas Marx, and Martin Scheffner. (2021) 2021. “Structural And Functional Consequences Of Nedd8 Phosphorylation.”. Nature Communications 12 (1): 5939. doi:10.1038/s41467-021-26189-9.
RS/tRNA Pair Development Year
2015
ncAA(s) Incorporated
phosphoserine
ncAA Structure (png, jpg, jpeg)
ncAA Utility
Used to produce site-specifically phosphorylated proteins at a serine residue via genetic code expansion versus other protein phosphorylation strategies.
2-amino-4-phosphonobutanoic acid (nhpSer)
ncAA Structure (png, jpg, jpeg)
ncAA Utility
Used to produce proteins that mimic site-specific phosphorylation at a serine residue - where the phosphate group cannot by hydrolyzed - via genetic code expansion versus other protein phosphorylation strategies.
RS Organism of Origin
Parent RS
RS Mutations
E412P
E414F
P495M
I496W
F529S
E414F
P495M
I496W
F529S
tRNA Organism of Origin
Parent tRNA
tRNA Anticodon
CUA
Other tRNA Mutations
G29A
A31U
G34C
C35
G37A
U39A
C40U
C41U
A31U
G34C
C35
G37A
U39A
C40U
C41U
RS/tRNA Availability
Component Addgene IDs are #173897 for SepRS2/Sep-tRNA(B4) machinery, and #174075 and #174076 for sfGFP WT and 150TAG constructs, respectively.
E. coli strains:
BL21(DE3) ΔserB - Addgene #34929
B95(DE3) ΔA ΔfabR ΔserB - Addgene #197655
Keio ΔserC - Obtained from GE-Dharmacon
The sfGFP/POI plasmids are not the ones originally used with this system, but work well with it. You can replace the sfGFP constructs by cloning in another POI.
E. coli strains:
BL21(DE3) ΔserB - Addgene #34929
B95(DE3) ΔA ΔfabR ΔserB - Addgene #197655
Keio ΔserC - Obtained from GE-Dharmacon
The sfGFP/POI plasmids are not the ones originally used with this system, but work well with it. You can replace the sfGFP constructs by cloning in another POI.
Used in what cell line?