Position-specific incorporation of a highly photodurable and blue-laser excitable fluorescent amino acid into proteins for fluorescence sensing.
A new fluorescent amino acid, L-2-acridonylalanine, was incorporated into proteins at specific positions using 4-base codon/anticodon strategy. The efficiency of the incorporation was high enough to obtain enough quantities of the mutants. The acridonyl group was highly fluorescent when it was excited at the wavelengths of blue-lasers and was highly photodurable compared with conventional fluorophores often used for biological analyses. The fluorescence intensity was sensitive to small changes in the polarity of the environment. When the nonnatural amino acid was incorporated into specific positions of streptavidin, the mutant protein worked as a fluorescent sensor to biotin. Similarly, when the amino acid was incorporated into camel single-chain antibody, the mutant protein sensitively responded to the antigen molecule. The high incorporation efficiency, the high photodurability, the excitability with blue-lasers, and high sensitivity to the environment make the acridonylalanine as the promising fluorescent amino acid for sensing small molecules when incorporated into specific positions of various antibodies, receptors, and enzymes.
Hamada
Hiroyuki
H
Department of Bioscience and Biotechnology, Faculty of Engineering, Okayama University, 3-1-1 Tsushimanaka, Okayama 700-8530, Japan.
Kameshima
Naoko
N
Szymańska
Aneta
A
Wegner
Katarzyna
K
Lankiewicz
Łeszek
Ł
Shinohara
Hiroaki
H
Taki
Masumi
M
Sisido
Masahiko
M
D000166:Acridines / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D054831:Acridones; D000596:Amino Acids / Q000138:chemical synthesis* / Q000737:chemistry* / Q000378:metabolism; D000818:Animals; D002162:Camelus; D002645:Chickens; D005456:Fluorescent Dyes / Q000138:chemical synthesis* / Q000737:chemistry*; D007074:Immunoglobulin G / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007202:Indicators and Reagents / Q000737:chemistry / Q000494:pharmacology; D007834:Lasers; D008027:Light; D008958:Models, Molecular; D009113:Muramidase / Q000737:chemistry / Q000378:metabolism; D009154:Mutation; D013050:Spectrometry, Fluorescence; D019809:Streptavidin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D000166:Acridines; D054831:Acridones; D000596:Amino Acids; D005456:Fluorescent Dyes; D007074:Immunoglobulin G; D007202:Indicators and Reagents; C041300:acridone; D019809:Streptavidin; D009113:Muramidase