Protein reconstitution methods for visualizing biomolecular function in living cells.
One of the most challenging researches in current biology and medicinal chemistry is to understand how individual cellular molecules interact together in living cells. To visualize such molecules, genetically-encoded reporters have been used widely. The most common reporters are firefly luciferase, renilla luciferase, green fluorescent protein (GFP) and its variants with various spectral properties. In this review, novel design of split GFP and split luciferase is described. The principle is based on reconstitution of the split-reporter fragments when they are brought together into close proximity. The reconstitution methods are used for screening organelle-localized proteins, imaging dynamics of nuclear proteins and mRNAs in living cells, and visualizing protease activities in living animals. These methods are generally applicable for imaging of complex cellular processes and evaluating chemical effects in living cells and animals.
Ozawa
Takeaki
T
Department of Chemistry, School of Science, The University of Tokyo, Tokyo, Japan. ozawa@chem.s.u-tokyo.ac.jp
D000818:Animals; D015374:Biosensing Techniques / Q000379:methods*; D053148:Caspase 3 / Q000378:metabolism; D003952:Diagnostic Imaging / Q000379:methods*; D015195:Drug Design; D015723:Gene Library; D049452:Green Fluorescent Proteins; D049409:Luciferases, Firefly; D049410:Luciferases, Renilla; D051379:Mice; D008967:Molecular Biology / Q000379:methods*; D021381:Protein Transport; D012333:RNA, Messenger
D012333:RNA, Messenger; D049452:Green Fluorescent Proteins; D049410:Luciferases, Renilla; D049409:Luciferases, Firefly; D053148:Caspase 3