Effective photoregulation of gene expression by photo responsive T7 promoter.
We constructed a photoresponsive T7 promoter by tethering two 2,6-dimethyl azobenzenes (4-(2,6-dimethyl-phenlylazo)-benzoic acids) via D-threoninol linkers. Under UV light irradiation, the rate of transcription with T7 RNA polymerase (RNAP) on the photoresponsive promoter was 10-fold faster than that under visible light irradiation. Kinetic analysis revealed that K(m) of cis-cis-promoter (both of the introduced azobenzenes are in cis-form) with T7 RNAP was more than 60 times larger than that of trans-trans-form, indicating that the high photoregulatory efficiency was obtained mainly due to the remarkable difference in their affinity with RNAP. By attaching a photoresponsive promoter to the GFP gene, we also showed that photoregulation of gene expression became practicable.
Liang
Xingguo
X
Department of Molecular Design and Engineering, Graduate School of Engineering, Nagoya University, Chikusa, Nagoya 464-8603, Japan.
Wakuda
Ryuji
R
Tsuda
Yuichiro
Y
Nishioka
Hidenori
H
Asanuma
Hiroyuki
H
D001391:Azo Compounds / Q000737:chemistry*; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000528:radiation effects*; D007700:Kinetics; D011401:Promoter Regions, Genetic; D014158:Transcription, Genetic / Q000528:radiation effects*; D014466:Ultraviolet Rays; D014764:Viral Proteins / Q000378:metabolism*
C525945:2',6'-dimethylazobenzene; D001391:Azo Compounds; D014764:Viral Proteins; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases