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Title:
METHOD FOR CONSTRUCTING RECOMBINANT BACTERIUM FOR PRODUCING NON-NATIVE PROTEIN, AND UTILIZATION OF SAME
Number:
US20130095524A1
Publication Date:
2013-04-18
Location:
JP
Priority Date:
IPC clasifications:
C12N 15/70
Assignee:
RIKEN
Inventor:
YOKOYAMA SHIGEYUKI|MUKAI TAKAHITO|SAKAMOTO KENSAKU|MATSUMOTO AKIKO
Class / subclass:
Enabling Technologies
Industry:
Research
Abstract:

The present invention provides a novel method of producing a recombinant bacterium for production of a non-natural protein, including: (1) expressing tRNA in a bacterium, which tRNA recognizes UAG codon; (2) expressing an aminoacyl-tRNA synthetase in the bacterium, which aminoacyl-tRNA synthetase acylates the tRNA with a non-natural amino acid or an alpha-hydroxy acid; (3) (i) introducing a DNA construct into the bacterium, which DNA construct is for expressing, in the absence of a release factor for terminating translation at UAG codon, a function of at least one gene selected from the group consisting of genes each of which loses its function when a gene that codes for the release factor is defective and/or introducing an alteration into said at least one gene in a chromosome of the bacterium, which alteration is for expressing the function of said at least one gene in the absence of the release factor; and (4) causing the gene that codes for the release factor in the bacterium to be defective.

Claims:

A method of producing a recombinant bacterium for production of a non-natural protein, comprising the steps of: (1) expressing tRNA in a bacterium, which tRNA recognizes UAG codon; (2) expressing an aminoacyl-tRNA synthetase in a bacterium, which aminoacyl-tRNA synthetase acylates the tRNA with a non-natural amino acid or an α-hydroxy acid; (3) subjecting a bacterium to a process for expressing, in the absence of a release factor for terminating translation at UAG codon, a function of at least one gene selected from the group consisting of genes each of which loses its function when a gene that codes for the release factor is defective; and (4) causing the gene that codes for the release factor in a bacterium to be defective, the bacteria in the steps (1) through (4) being identical, and the process being (i) a process of introducing, into the bacterium, a DNA construct for expressing the function of said at least one gene in the absence of the release factor and/or (ii) a process of introducing, into said at least one gene in a chromosome of the bacterium, an alteration for expressing the function of said at least one gene in the absence of the release factor.

Patent Family:
EP2584037A1 US20130095524A1 WO2011158895A1