When: August 21, 2011 12:00
Where: Physics 377
Presenter: Jorge Verdin Ramos (Benjamin Podbilewicz lab, biology)
Link to paper: http://www.nature.com/nature/journal/v464/n7287/full/nature08817.html
Medical Research Council Laboratory of Molecular Biology, Hills Road, Cambridge
The in vivo, genetically programmed incorporation of designer amino acids allows the properties of proteins to be tailored with molecular precision1. The Methanococcus jannaschii tyrosyl-transfer-RNA synthetase–tRNACUA (MjTyrRS–tRNACUA)2, 3 and the Methanosarcina barkeri pyrrolysyl-tRNA synthetase–tRNACUA (MbPylRS–tRNACUA)4, 5, 6orthogonal pairs have been evolved to incorporate a range of unnatural amino acids in response to the amber codon in Escherichia coli1, 6, 7. However, the potential of synthetic genetic code expansion is generally limited to the low efficiency incorporation of a single type of unnatural amino acid at a time, because every triplet codon in the universal genetic code is used in encoding the synthesis of the proteome. To encode efficiently many distinct unnatural amino acids into proteins we require blank codons and mutually orthogonal aminoacyl-tRNA synthetase–tRNA pairs that recognize unnatural amino acids and decode the new codons. Here we synthetically evolve an orthogonal ribosome8, 9 (ribo-Q1) that efficiently decodes a series of quadruplet codons and the amber codon, providing several blank codons on an orthogonal messenger RNA, which it specifically translates8. By creating mutually orthogonal aminoacyl-tRNA synthetase–tRNA pairs and combining them with ribo-Q1 we direct the incorporation of distinct unnatural amino acids in response to two of the new blank codons on the orthogonal mRNA. Using this code, we genetically direct the formation of a specific, redox-insensitive, nanoscale protein cross-link by the bio-orthogonal cycloaddition of encoded azide- and alkyne-containing amino acids10. Because the synthetase–tRNA pairs used have been evolved to incorporate numerous unnatural amino acids1, 6, 7, it will be possible to encode more than 200 unnatural amino acid combinations using this approach. As ribo-Q1 independently decodes a series of quadruplet codons, this work provides foundational technologies for the encoded synthesis and synthetic evolution of unnatural polymers in cells.
Where: Physics 377
Presenter: Jorge Verdin Ramos (Benjamin Podbilewicz lab, biology)
Link to paper: http://www.nature.com/nature/journal/v464/n7287/full/nature08817.html
Encoding multiple unnatural amino acids via evolution of a quadruplet-decoding ribosome
Heinz Neumann, Kaihang Wang, Lloyd Davis, Maria Garcia-Alai & Jason W. ChinMedical Research Council Laboratory of Molecular Biology, Hills Road, Cambridge
The in vivo, genetically programmed incorporation of designer amino acids allows the properties of proteins to be tailored with molecular precision1. The Methanococcus jannaschii tyrosyl-transfer-RNA synthetase–tRNACUA (MjTyrRS–tRNACUA)2, 3 and the Methanosarcina barkeri pyrrolysyl-tRNA synthetase–tRNACUA (MbPylRS–tRNACUA)4, 5, 6orthogonal pairs have been evolved to incorporate a range of unnatural amino acids in response to the amber codon in Escherichia coli1, 6, 7. However, the potential of synthetic genetic code expansion is generally limited to the low efficiency incorporation of a single type of unnatural amino acid at a time, because every triplet codon in the universal genetic code is used in encoding the synthesis of the proteome. To encode efficiently many distinct unnatural amino acids into proteins we require blank codons and mutually orthogonal aminoacyl-tRNA synthetase–tRNA pairs that recognize unnatural amino acids and decode the new codons. Here we synthetically evolve an orthogonal ribosome8, 9 (ribo-Q1) that efficiently decodes a series of quadruplet codons and the amber codon, providing several blank codons on an orthogonal messenger RNA, which it specifically translates8. By creating mutually orthogonal aminoacyl-tRNA synthetase–tRNA pairs and combining them with ribo-Q1 we direct the incorporation of distinct unnatural amino acids in response to two of the new blank codons on the orthogonal mRNA. Using this code, we genetically direct the formation of a specific, redox-insensitive, nanoscale protein cross-link by the bio-orthogonal cycloaddition of encoded azide- and alkyne-containing amino acids10. Because the synthetase–tRNA pairs used have been evolved to incorporate numerous unnatural amino acids1, 6, 7, it will be possible to encode more than 200 unnatural amino acid combinations using this approach. As ribo-Q1 independently decodes a series of quadruplet codons, this work provides foundational technologies for the encoded synthesis and synthetic evolution of unnatural polymers in cells.
No comments:
Post a Comment