Prevention of incorporation of non-standard amino acids into...

C - Chemistry – Metallurgy – 12 – P

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C12P 21/02 (2006.01) C12N 9/06 (2006.01) C12N 15/53 (2006.01)

Patent

CA 2539996

The instant invention is drawn to the methods and compositions necessary to provide recombinant proteins with a substantially reduced or eliminated content of norleucine or other non-standard amino acids. Various embodiments of the invention provide for the substantial elimination of the incorporation of non-standard amino acids into recombinant proteins by the co-expression or enhanced expression of a protein (or the enzymatically active portion thereof) capable of degrading norleucine or other non-standard amino acids, including norvaline, beta-methylnorleucine, and homoisoleucine. In certain particular embodiments of the invention, the norleucine is degraded by a glutamate dehydrogenase, a leucine dehydrogenase, a valine dehydrogenase, a phenylalanine dehydrogenase, a glutamate/leucine/phenylalanine/valine dehydrogenase, or an opine dehydrogenase. Also provided are the cells and DNA constructs for carrying out these methods.

La présente invention se rapporte à des procédés et à des compositions nécessaires à la génération de protéines recombinées dont la teneur en norleucine ou en d'autres acides aminés non standard est sensiblement réduite ou nulle. Divers modes de réalisation de l'invention concernent la quasi-élimination de l'incorporation d'acides aminés non standard dans des protéines recombinées, par la coexpression ou l'expression amplifiée d'une protéine (ou de la partie enzymatiquement active de cette dernière) capable de dégrader la norleucine ou d'autres acides aminés non standard, notamment la norvaline, la bétaméthylnorleucine et l'homo-isoleucine. Dans certains modes de réalisation particuliers de l'invention, la norleucine est dégradée par une glutamate déshydrogénase, une leucine déshydrogénase, une valine déshydrogénase, une phénylalanine déshydrogénase, une glutamate/leucine/phénylalanine/valine déshydrogénase, ou une opine déshydrogénase. L'invention a également trait à des cellules et à des ADN de synthèse permettant de mettre en oeuvre lesdits procédés.

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