Abstract
Optical imaging of protein–protein interactions (PPIs) facilitates comprehensive elucidation of intracellular molecular events. The present protocol demonstrates an optical measure for visualizing molecular tension triggered by any PPI in mammalian cells. A unique design of single-chain probes was fabricated, in which a full-length artificial luciferase (ALuc ®) was sandwiched between two model proteins of interest, e.g.,FKBP and FRB. A molecular tension probe comprising ALuc23 greatly enhances the bioluminescence in response to varying concentrations of rapamycin, and named “tension probe (TP)”. The basic probe design can be further modified towards eliminating the C-terminal end of ALuc and was found to improve signal-to-background ratios, named “combinational probe”. TPs may become an important addition to the tool box of bioassays in the determination of protein dynamics of interest in mammalian cells.
| Original language | English |
|---|---|
| Title of host publication | Methods in Molecular Biology |
| Publisher | Humana Press Inc. |
| Pages | 183-193 |
| Number of pages | 11 |
| Volume | 1461 |
| DOIs | |
| Publication status | Published - 2016 |
| Externally published | Yes |
Publication series
| Name | Methods in Molecular Biology |
|---|---|
| Volume | 1461 |
| ISSN (Print) | 10643745 |
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Keywords
- Bioluminescence
- Bioluminescent imaging
- Combinational probe
- Protein–protein interactions
- Tension probe
ASJC Scopus subject areas
- Molecular Biology
- Genetics
Cite this
An Aluc-based molecular tension probe for sensing intramolecular protein–Protein interactions. / Kim, Sung Bae; Nishihara, Ryo; Suzuki, Koji.
Methods in Molecular Biology. Vol. 1461 Humana Press Inc., 2016. p. 183-193 (Methods in Molecular Biology; Vol. 1461).Research output: Chapter in Book/Report/Conference proceeding › Chapter
}
TY - CHAP
T1 - An Aluc-based molecular tension probe for sensing intramolecular protein–Protein interactions
AU - Kim, Sung Bae
AU - Nishihara, Ryo
AU - Suzuki, Koji
PY - 2016
Y1 - 2016
N2 - Optical imaging of protein–protein interactions (PPIs) facilitates comprehensive elucidation of intracellular molecular events. The present protocol demonstrates an optical measure for visualizing molecular tension triggered by any PPI in mammalian cells. A unique design of single-chain probes was fabricated, in which a full-length artificial luciferase (ALuc ®) was sandwiched between two model proteins of interest, e.g.,FKBP and FRB. A molecular tension probe comprising ALuc23 greatly enhances the bioluminescence in response to varying concentrations of rapamycin, and named “tension probe (TP)”. The basic probe design can be further modified towards eliminating the C-terminal end of ALuc and was found to improve signal-to-background ratios, named “combinational probe”. TPs may become an important addition to the tool box of bioassays in the determination of protein dynamics of interest in mammalian cells.
AB - Optical imaging of protein–protein interactions (PPIs) facilitates comprehensive elucidation of intracellular molecular events. The present protocol demonstrates an optical measure for visualizing molecular tension triggered by any PPI in mammalian cells. A unique design of single-chain probes was fabricated, in which a full-length artificial luciferase (ALuc ®) was sandwiched between two model proteins of interest, e.g.,FKBP and FRB. A molecular tension probe comprising ALuc23 greatly enhances the bioluminescence in response to varying concentrations of rapamycin, and named “tension probe (TP)”. The basic probe design can be further modified towards eliminating the C-terminal end of ALuc and was found to improve signal-to-background ratios, named “combinational probe”. TPs may become an important addition to the tool box of bioassays in the determination of protein dynamics of interest in mammalian cells.
KW - Bioluminescence
KW - Bioluminescent imaging
KW - Combinational probe
KW - Protein–protein interactions
KW - Tension probe
UR - http://www.scopus.com/inward/record.url?scp=84978764170&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84978764170&partnerID=8YFLogxK
U2 - 10.1007/978-1-4939-3813-1_15
DO - 10.1007/978-1-4939-3813-1_15
M3 - Chapter
AN - SCOPUS:84978764170
VL - 1461
T3 - Methods in Molecular Biology
SP - 183
EP - 193
BT - Methods in Molecular Biology
PB - Humana Press Inc.
ER -