Split Gaussia luciferase-based bioluminescence template for tracing protein dynamics in living cells.
The goal of the present study is to develop a small luminescence template, in which any protein may be incorporated, for illuminating the dynamics of protein signaling. We first determined optimal fragmentation sites of Gaussia princeps-derived luciferase (GLuc). The utility of the template was demonstrated with calmodulin (CaM) and a peptide-linked CaM, which were sandwiched between the fragments of split GLuc dissected at Q105. Living mammalian cells with the probe quickly emitted luminescence in response to endo- and exogenous Ca2+ levels. The applicability of the template was expanded to the visualization of the phosphorylation of the estrogen receptor and interaction of the androgen receptor with a peptide via an intramolecular complementation between GLuc fragments. It also provides the smallest bioluminescence template ever synthesized. Considering that conformational changes of proteins generally occur for signal transductions, the present luminescence template will contribute to the exploration of intracellular molecular events with signaling proteins.
Kim
Sung Bae
SB
Research Institute for Environmental Management Technology, National Institute of Advanced Industrial Science and Technology (AIST), 16-1 Onogawa, Tsukuba 305-8569, Japan.
Sato
Moritoshi
M
Tao
Hiroaki
H
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002118:Calcium / Q000737:chemistry / Q000378:metabolism; D002147:Calmodulin / Q000737:chemistry / Q000378:metabolism; D002522:Chlorocebus aethiops; D033342:Copepoda / Q000201:enzymology; D008024:Ligands; D008156:Luciferases / Q000737:chemistry* / Q000378:metabolism; D008163:Luminescent Measurements / Q000379:methods*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D011506:Proteins / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D011960:Receptors, Estrogen / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D011965:Receptors, Glucocorticoid / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D011980:Receptors, Progesterone / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D015290:Second Messenger Systems; D014982:Xenopus laevis
D002147:Calmodulin; D008024:Ligands; D010446:Peptide Fragments; D011506:Proteins; D011960:Receptors, Estrogen; D011965:Receptors, Glucocorticoid; D011980:Receptors, Progesterone; D008156:Luciferases; D002118:Calcium