Cloning, sequencing and expression of a comamonas...

C - Chemistry – Metallurgy – 12 – N

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C12N 15/53 (2006.01) C12N 1/20 (2006.01) C12N 9/02 (2006.01)

Patent

CA 2418155

Cyclopentanone 1,2-monooxygenase (CPMO) from Comamonas (previously Pseudomonas) sp. strain NCIMB 9872 carries out the second step of a degradation pathway that allows the bacterium to use cyclopentanol as a sole carbon source for growth. In the present invention there is reported the localization of the CPMO-encoding gene (cpnB) on a 4.3-kb SphI fragment, the determination of its sequence. The 550-amino acid CPMO polypeptide (Mr, 62,111) encoded by the gene was found to have 36.5% identity with the sequence of cyclohexanone 1,2-monooxygenase (CHMO) of Acinetobacter sp. strain NCIMB 9871. The 62-kDa CPMO was expressed in E. coli as an IPTG-inducible protein.

Le cyclopentanone 1,2-monooxygénase (CPMO) du Comamonas (antérieurement Pseudomonas) sp. souche NCIMB 9872 exécute la deuxième étape d'un mécanisme de dégradation permettant à la bactérie d'utiliser du carbone comme source unique pour sa croissance. L'invention porte sur la localisation du gène (cpnB) codant pour le CPMO, sur un fragment SphI de 4,3-kb, et la détermination de sa séquence. On a découvert que le polypeptide (<I>M</I>¿r?, 62,111) de l'acide aminé 550 du CPMO codé par ledit gène présentait 36,5 % de similitude avec la séquence de cyclohexanone 1,2-monooxygénase (CHMO) de l'Acinetobacter sp. souche NCIMB 9871. Le CPMO de 62-kDa à été exprimé dans le E. coli en tant que protéine inductible par l'IPTG.

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