Position-specific incorporation of dansylated non-natural amino acids into streptavidin by using a four-base codon.
Novel non-natural amino acids carrying a dansyl fluorescent group were designed, synthesized, and incorporated into various positions of streptavidin by using a CGGG four-base codon in an Escherichia coli in vitro translation system. 2,6-Dansyl-aminophenylalanine (2,6-dnsAF) was found to be incorporated into the protein more efficiently than 1,5-dansyl-lysine, 2,6-dansyl-lysine, and 1,5-dansyl-aminophenylalanine. Fluorescence measurements indicate that the position-specific incorporation of the 2,6-dnsAF is a useful technique to probe protein structures. These results also indicate that well-designed non-natural amino acids carrying relatively large side chains can be accepted as substrates of the translation system.
Hohsaka
Takahiro
T
Department of Bioscience and Biotechnology, Okayama University, 3-1-1 Tsushimanaka, Okayama 700-8530, Japan. hohsaka@jaist.ac.jp
Muranaka
Norihito
N
Komiyama
Chie
C
Matsui
Kinue
K
Takaura
Satomi
S
Abe
Ryoji
R
Murakami
Hiroshi
H
Sisido
Masahiko
M
D000596:Amino Acids / Q000138:chemical synthesis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D003062:Codon; D004926:Escherichia coli / Q000235:genetics; D005240:Feasibility Studies; D005456:Fluorescent Dyes / Q000737:chemistry*; D007202:Indicators and Reagents / Q000737:chemistry*; D008958:Models, Molecular; D015394:Molecular Structure; D017354:Point Mutation; D014176:Protein Biosynthesis; D015202:Protein Engineering; D017433:Protein Structure, Secondary; D012313:RNA / Q000737:chemistry; D012333:RNA, Messenger / Q000235:genetics; D013050:Spectrometry, Fluorescence; D019809:Streptavidin / Q000737:chemistry*; D013379:Substrate Specificity
D000596:Amino Acids; D003062:Codon; D005456:Fluorescent Dyes; D007202:Indicators and Reagents; D012333:RNA, Messenger; D012313:RNA; D019809:Streptavidin