Cell-free translation reconstituted with purified components.
We have developed a protein-synthesizing system reconstituted from recombinant tagged protein factors purified to homogeneity. The system was able to produce protein at a rate of about 160 microg/ml/h in a batch mode without the need for any supplementary apparatus. The protein products were easily purified within 1 h using affinity chromatography to remove the tagged protein factors. Moreover, omission of a release factor allowed efficient incorporation of an unnatural amino acid using suppressor transfer RNA (tRNA).
Shimizu
Y
Y
Department of Chemistry and Biotechnology, Graduate School of Engineering, The University of Tokyo, Bldg FSB401, 5-1-5, Kasiwanoha, Kashiwa, Chiba prefecture 277-8562, Japan.
Inoue
A
A
Tomari
Y
Y
Suzuki
T
T
Yokogawa
T
T
Nishikawa
K
K
Ueda
T
T
D000596:Amino Acids / Q000737:chemistry; D002474:Cell-Free System; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D006639:Histidine / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D014176:Protein Biosynthesis; D012343:RNA, Transfer / Q000378:metabolism; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism; D013997:Time Factors
D000596:Amino Acids; D012346:RNA, Transfer, Amino Acyl; D011994:Recombinant Proteins; D006639:Histidine; D012343:RNA, Transfer