Methods for producing hyaluronan in a recombinant host cell

C - Chemistry – Metallurgy – 12 – N

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Details

C12N 15/62 (2006.01) C12N 1/21 (2006.01) C12N 9/04 (2006.01) C12N 9/10 (2006.01) C12N 15/52 (2006.01) C12N 15/53 (2006.01) C12N 15/54 (2006.01) C12N 15/61 (2006.01) C12P 19/04 (2006.01) C12P 19/26 (2006.01) C12P 21/02 (2006.01)

Patent

CA 2471148

The present invention relates to methods for producing a hyaluronic acid, comprising: (a) cultivating a Bacillus host cell under conditions suitable for production of the hyaluronic acid, wherein the Bacillus host cell comprises a nucleic acid construct comprising a hyaluronan synthase encoding sequence operably linked to a promoter sequence foreign to the hyaluronan synthase encoding sequence; and (b) recovering the hyaluronic acid from the cultivation medium. The present invention also relates to an isolated nucleic acid sequence encoding a hyaluronan synthase operon comprising a hyaluronan synthase gene and a UDP-glucose 6-dehydrogenase gene, and optionally one or more genes selected from the group consisting of a UDP-glucose pyrophosphorylase gene, UDP-N-acetylglucosamine pyrophosphorylase gene, and glucose-6-phosphate isomerase gene. The present invention also relates to isolated nucleic acid sequences encoding a UDP-glucose 6-dehydrogenase, UDP- glucose pyrophosphorylase, and UDP-N-acetylglucosamine pyrophosphorylase.

La présente invention concerne des procédés de production d'un acide hyaluronique, consistant à : (a) cultiver une cellule hôte de Bacillus dans des conditions adaptées à la production de l'acide hyaluronique, ladite cellule hôte de Bacillus comprenant une construction d'acide nucléique comportant une séquence codant pour la hyaluronane synthase fonctionnellement liée à une séquence promotrice étrangère à la séquence codant pour la hyaluronane synthase ; et (b) récupérer l'acide hyaluronique à partir du milieu de culture. L'invention concerne également une séquence d'acide nucléique isolée codant pour un opéron de la hyaluronane synthase comprenant un gène de hyaluronane synthase et un gène d'UDP-glucose 6-déshydrogénase, ainsi qu'éventuellement un ou plusieurs gènes choisis dans le groupe comprenant un gène d'UDP-glucose pyrophosphorylase, un gène d'UDP-N-acétylglucosamine pyrophosphorylase et un gène de glucose-6-phosphate isomérase. L'invention concerne par ailleurs des séquences d'acide nucléique isolées codant pour une UDP-glucose 6-déshydrogénase, une UDP-glucose pyrophosphorylase et une UDP-N-acétylglucosamine pyrophosphorylase.

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