Mammalian expression of infrared fluorescent proteins engineered from a bacterial phytochrome.
Visibly fluorescent proteins (FPs) from jellyfish and corals have revolutionized many areas of molecular and cell biology, but the use of FPs in intact animals, such as mice, has been handicapped by poor penetration of excitation light. We now show that a bacteriophytochrome from Deinococcus radiodurans, incorporating biliverdin as the chromophore, can be engineered into monomeric, infrared-fluorescent proteins (IFPs), with excitation and emission maxima of 684 and 708 nm, respectively; extinction coefficient >90,000 M(-1) cm(-1); and quantum yield of 0.07. IFPs express well in mammalian cells and mice and spontaneously incorporate biliverdin, which is ubiquitous as the initial intermediate in heme catabolism but has negligible fluorescence by itself. Because their wavelengths penetrate tissue well, IFPs are suitable for whole-body imaging. The IFPs developed here provide a scaffold for further engineering.
Shu
Xiaokun
X
Howard Hughes Medical Institute, University of California at San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0647, USA.
Royant
Antoine
A
Lin
Michael Z
MZ
Aguilera
Todd A
TA
Lev-Ram
Varda
V
Steinbach
Paul A
PA
Tsien
Roger Y
RY
D000256:Adenoviridae / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D001664:Biliverdine / Q000737:chemistry / Q000378:metabolism; D002460:Cell Line; D034301:Deinococcus / Q000737:chemistry*; D003952:Diagnostic Imaging; D005453:Fluorescence; D006801:Humans; D008099:Liver / Q000033:anatomy & histology; D008164:Luminescent Proteins / Q000737:chemistry / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D010834:Phytochrome / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015202:Protein Engineering; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D013055:Spectrophotometry, Infrared; D051598:Whole Body Imaging
D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D010834:Phytochrome; D001664:Biliverdine