PMID- 21900305
OWN - NLM
STAT- MEDLINE
DCOM- 20120215
LR  - 20211203
IS  - 1741-0134 (Electronic)
IS  - 1741-0126 (Print)
IS  - 1741-0126 (Linking)
VI  - 24
IP  - 11
DP  - 2011 Nov
TI  - An evaluation tool for FKBP12-dependent and -independent mTOR inhibitors using a 
      combination of FKBP-mTOR fusion protein, DSC and NMR.
PG  - 811-7
LID - 10.1093/protein/gzr045 [doi]
AB  - Mammalian target of rapamycin (mTOR), a large multidomain protein kinase, 
      regulates cell growth and metabolism in response to environmental signals. The 
      FKBP rapamycin-binding (FRB) domain of mTOR is a validated therapeutic target for 
      the development of immunosuppressant and anticancer drugs but is labile and 
      insoluble. Here we designed a fusion protein between FKBP12 and the FRB domain of 
      mTOR. The fusion protein was successfully expressed in Escherichia coli as a 
      soluble form, and was purified by a simple two-step chromatographic procedure. 
      The fusion protein exhibited increased solubility and stability compared with the 
      isolated FRB domain, and facilitated the analysis of rapamycin and FK506 binding 
      using differential scanning calorimetry (DSC) and solution nuclear magnetic 
      resonance (NMR). DSC enabled the rapid observation of protein-drug interactions 
      at the domain level, while NMR gave insights into the protein-drug interactions 
      at the residue level. The use of the FKBP12-FRB fusion protein combined with DSC 
      and NMR provides a useful tool for the efficient screening of FKBP12-dependent as 
      well as -independent inhibitors of the mTOR FRB domain.
FAU - Sekiguchi, Mitsuhiro
AU  - Sekiguchi M
AD  - Analysis and Pharmacokinetics Research Labs, Department of Drug Discovery, 
      Astellas Pharma Inc., Tsukuba 350-8585, Japan.
FAU - Kobashigawa, Yoshihiro
AU  - Kobashigawa Y
FAU - Kawasaki, Masashi
AU  - Kawasaki M
FAU - Yokochi, Masashi
AU  - Yokochi M
FAU - Kiso, Tetsuo
AU  - Kiso T
FAU - Suzumura, Ken-ichi
AU  - Suzumura K
FAU - Mori, Keitaro
AU  - Mori K
FAU - Teramura, Toshio
AU  - Teramura T
FAU - Inagaki, Fuyuhiko
AU  - Inagaki F
LA  - eng
PT  - Evaluation Study
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20110906
PL  - England
TA  - Protein Eng Des Sel
JT  - Protein engineering, design & selection : PEDS
JID - 101186484
RN  - 0 (Enzyme Inhibitors)
RN  - 0 (Ligands)
RN  - 0 (Recombinant Fusion Proteins)
RN  - EC 2.7.1.1 (MTOR protein, human)
RN  - EC 2.7.11.1 (TOR Serine-Threonine Kinases)
RN  - EC 5.2.1.- (Tacrolimus Binding Protein 1A)
SB  - IM
MH  - Calorimetry, Differential Scanning/*methods
MH  - Enzyme Inhibitors/chemistry/*pharmacology
MH  - Humans
MH  - Ligands
MH  - Magnetic Resonance Spectroscopy/*methods
MH  - Protein Binding/drug effects
MH  - Protein Structure, Tertiary
MH  - Recombinant Fusion Proteins/chemistry/*metabolism
MH  - TOR Serine-Threonine Kinases/antagonists & inhibitors/chemistry/*metabolism
MH  - Tacrolimus Binding Protein 1A/antagonists & inhibitors/chemistry/*metabolism
PMC - PMC3196870
EDAT- 2011/09/09 06:00
MHDA- 2012/02/16 06:00
PMCR- 2011/09/06
CRDT- 2011/09/09 06:00
PHST- 2011/09/09 06:00 [entrez]
PHST- 2011/09/09 06:00 [pubmed]
PHST- 2012/02/16 06:00 [medline]
PHST- 2011/09/06 00:00 [pmc-release]
AID - gzr045 [pii]
AID - 10.1093/protein/gzr045 [doi]
PST - ppublish
SO  - Protein Eng Des Sel. 2011 Nov;24(11):811-7. doi: 10.1093/protein/gzr045. Epub 
      2011 Sep 6.