RNA aptamer binding to polyhistidine-tag.
Polyhistidine-tag (His-tag) is a powerful tool for purification of recombinant protein. His-tagged protein can be affinity-purified by using resins immobilizing Ni2+ or anti-His-tag antibodies. However, Ni2+-affinity-purification is prevented by the presence of divalent cations. The purification with antibodies has contamination of antibody peptides, which interferes with following analysis. In the present study, we isolated RNA aptamers binding to His-tag. The best clone, named shot47, bound to the target with low picomolar dissociation constant. In the presence of divalent cations, shot47 was substitutable for antibodies against His-tag on ELISA, immunoprecipitation, and Western blotting. Shot47 can be synthesized easily by in vitro transcription. Thus, shot47 would be applicable as a useful and cost-effective tool for biochemical analyses.
Tsuji
Shoutaro
S
Division of Cancer Therapy, Kanagawa Cancer Center Research Institute, Yokohama-shi, Kanagawa 241-0815, Japan. stsuji@gancen.asahi.yokohama.jp
Tanaka
Taku
T
Hirabayashi
Naomi
N
Kato
Shintaro
S
Akitomi
Joe
J
Egashira
Hazuki
H
Waga
Iwao
I
Ohtsu
Takashi
T
D000906:Antibodies / Q000737:chemistry / Q000276:immunology; D052157:Aptamers, Nucleotide / Q000737:chemistry* / Q000302:isolation & purification; D001483:Base Sequence; D002846:Chromatography, Affinity; D006639:Histidine / Q000737:chemistry*; D008263:Macrophage Migration-Inhibitory Factors / Q000302:isolation & purification; D009690:Nucleic Acid Conformation; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification*; D012313:RNA / Q000737:chemistry* / Q000302:isolation & purification
D000906:Antibodies; D052157:Aptamers, Nucleotide; D008263:Macrophage Migration-Inhibitory Factors; D011506:Proteins; C033223:polyhistidine; D006639:Histidine; D012313:RNA