Amidases, nucleic acids encoding them and methods for making...

C - Chemistry – Metallurgy – 12 – N

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C12N 15/55 (2006.01) A01H 5/00 (2006.01) A01K 67/027 (2006.01) A23C 19/14 (2006.01) A61K 31/7088 (2006.01) A61K 38/46 (2006.01) A61K 48/00 (2006.01) C07H 21/00 (2006.01) C07K 16/40 (2006.01) C12N 9/78 (2006.01) C12N 9/80 (2006.01) C12N 11/00 (2006.01) C12N 15/09 (2006.01) C12N 15/10 (2006.01) C12N 15/63 (2006.01) C12P 13/02 (2006.01) C12P 19/34 (2006.01) C12P 21/02 (2006.01) C12P 35/00 (2006.01) C12P 35/06 (2006.01) C12P 41/00 (2006.01) C12Q 1/34 (2006.01) G01N 33/573 (2006.01) G06F 19/00 (2006.01) A61K 38/

Patent

CA 2474567

This invention provides amidases, polynucleotides encoding the amidases, methods of making and using these polynucleotides and polypeptides. In one aspect, the invention provides enzymes having secondary amidase activity, e.g., having activity in the hydrolysis of amides, including enzymes having peptidase, protease and/or hydantoinase activity. In alternative aspects, the enzymes of the invention can be used to used to increase flavor in food (e.g., enzyme ripened cheese), promote bacterial and fungal killing, modify and de- protect fine chemical intermediates, synthesize peptide bonds, carry out chiral resolutions, hydrolyze Cephalosporin C. The enzymes of the invention can be used to generate 7-aminocephalosporanic acid (7-ACA) and semi-synthetic cephalosporin antibiotics, including caphalothin, cephaloridine and cefuroxime. The enzymes of the invention can be used as antimicrobial agents, e.g., as cell wall hydrolytic agents. The invention also provides a fluorescent amidase substrate comprising 7-(.epsilon.-D-2- aminoadipoyladipoylamido)-4-methylcoumarin.

L'invention concerne des amidases, des polynucléotides codant lesdites amidases, ainsi que des procédés de fabrication et d'utilisation desdits polynucléotides et polypetides. Dans un aspect, l'invention concerne des enzymes présentant une activité amidase secondaire, par exemple une activité dans l'hydrolyse des amides, notamment des enzymes présentant une activité peptidase, protéase et/ou hydantoïnase. Dans d'autres aspects, les enzymes selon l'invention peuvent être utilisées pour augmenter la saveur d'aliments (des fromages affinés aux enzymes, par exemple), renforcer l'élimination des bactéries et des champignons, modifier et déprotéger des intermédiaires chimiques fins, synthétiser des liaisons peptidiques, réaliser des résolutions chirales, hydrolyser la Céphalosporine C. Les enzymes selon l'invention peuvent être utilisées pour produire de l'acide 7-aminocéphalosporanique (7-ACA) et des antibiotiques céphalosporine semi-synthétiques, notamment la caphalothine, la céphaloridine et la céfuroxime. Les enzymes selon l'invention peuvent également être utilisées en tant qu'agents antimicrobiens, par exemple en tant qu'agents hydrolytiques dégradant la paroi cellulaire. La présente invention concerne également un substrat amidase fluorescent contenant de la 7-(.epsilon.-D-2-aminoadipoylamido)-4-méthylcoumarine.

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