Roche Vitamins Inc.

Dsm Nutritional Products Inc.

Hoffmann-la Roche Inc.

Akira Asakura

Tatsuo Hoshino

Masako Shinjoh

Noribumi Tomiyama

Tatsuya Kiyasu

Yoshie Nagahashi

Setsuko Ojima

The present invention is directed to a novel cytochrome c oxidase complex, genetic materials useful for the preparation of the said complex, such as recombinant polypeptides involved in cytochrome c oxidase complex, recombinant DNA fragments, expression vectors, recombinant organisms and the like. Those novel cytochrome c oxidase complex and genetic materials may be originated from a microorganism having the identifying characteristics ofDSM 4025. The present invention also provides a method for the preparation of the said novel recombinant cytochrome c oxidase complex and a process for the production of 2-keto-L-gulonic acid (2KGA).

Cytochrome c oxidase enzyme complex

The present invention is directed to a novel cytochrome c oxidase complex, genetic materials useful for the preparation of the said complex, such as recombinant polypeptides involved in cytochrome c oxidase complex, recombinant DNA fragments, expression vectors, recombinant organisms and the like. Those novel cytochrome c oxidase complex and genetic materials may be originated from a microorganism having the identifying characteristics of Gluconobacter oxydans DSM 4025. The present invention also provides a method for the preparation of the said novel recombinant cytochrome c oxidase complex and a process for the production of 2-keto-L-gulonic acid (2KGA).

A process for producing L-ascorbic acid or D-erythorbic acid, or in each case its sodium, potassium or calcium salt, from 2-keto-L-gulonic acid or 2-keto-D-gluconic acid, or in each case its sodium, potassium or calcium salt, that involves incubating 2-keto-L-gulonic acid or 2-keto-D-gluconic acid, each as the free acid or as its sodium, potassium or calcium salt, and the cells of a thermoacidophilic microorganism at temperatures from about 30&deg; C. to about 100&deg; C., and at a pH from about 1 to about 6, in a solution, to form L-ascorbic acid or D-erythorbic acid or an appropriate salt thereof, and isolating said product from the solution.

Microbial process for producing L-ascorbic acid, D-erythorbic acid, and salts thereof

The present invention is directed to a recombinant enzymes having alcohol and aldehyde dehydrogenase activity which comprises one or mom recombinant polypeptides selected from the group consisting of polypeptides which are identified by SEQ ID NO 5, SEQ ID NO 6, SEQ ID NO 7, SEQ ID NO 8 and chimeric recombinant polypeptides that are a chimeric combination of at least two of the following amino acid sequences identified by SEQ ID NO 5, SEQ ID NO 6, SEQ ID NO 7, SEQ ID NO 8 and functional derivatives of the polypeptides identified above which contain addition, insertion, deletion and/or substitution of one or more amino acid residues, wherein said enzymatic polypeptides have said alcohol and aldehyde dehydrogenase activity. DNA molecules encoding the recombinant polypeptides, vectors comprising such DNA molecules, host cells transformed by such vectors and processes for the production of such recombinant enzymes are provided. Furthermore the recombinant enzymes having alcohol and aldehyde dehydrogenase activity are used for obtaining aldehydes, ketones or carboxylic acids, and specifically, 2-keto-L-gulonic acid an intermediate for the production of L-ascorbic acid (vitamin C).

Alcohol/aldehyde dehydrogenase

This invention is directed to cytochrome c polypeptides and polynucleotides encoding the polypeptides.

Cytochrome c and polynucleotides encoding cytochrome c

The present invention provides processes for making biotin from desthiobiotin by either contacting desthiobiotin with an enzyme reaction mixture containing bioB gene product and nifU gene product and/or nifS gene product and isolating the biotin or cultivating a microorganism transformed with DNA encoding the bioB gene product, nifU gene product and nifS gene product and isolating the biotin.

Production of biotin

A process for producing L-ascorbic acid (vitamin C) from 2-keto-L-gulonic acid or D-erythorbic acid from 2-keto-D-gluconic acid by contacting 2-keto-L-gulonic acid or 2-keto-D-gluconic acid, respectively, in solution with a lactonase, particularly one belonging to the enzyme class EC 3.1.1.x, according to the classification of Enzyme Nomenclature. The solvent for this reaction can be water, an aqueous alcohol, a non-alcoholic organic solvent or a mixture of an aqueous alcohol and a non-alcoholic organic solvent. The contacting is generally performed in a temperature range of 0.degree. C. to 120.degree. C. and a pH range of 1.5 to 12. In each case the starting material can be in the form of the free acid, the sodium salt, or the calcium salt. The so-produced vitamin C has very well known uses, and the alternatively produced D-erythorbic acid is useful as an antioxidant for food additives.

Manufacture of L-ascorbic acid and D-erythorbic acid

This invention is directed to cytochrome c.sub.551 polypeptides and polynucleotides encoding the polypeptides.

Cytochrome C and polynucleotides encoding cytochrome C

The present invention relates to a novel alcohol/aldehyde dehydrogenase (hereinafter referred to as AADH), a process for producing the same and a process for producing aldehydes, carboxylic acids and ketones, especially 2-keto-L-gulonic acid (hereinafter referred to as 2-KGA) utilizing said enzyme.

Gluconobacter alcohol/aldehyde dehydrogenase

The present invention relates to a novel alcohol/aldehyde dehydro-genase (hereinafter referred to as AADH), a process for producing the same and a process for producing aldehydes, carboxylic acids and ketones, especially 2-keto-L-gulonic acid (hereinafter referred to as 2-KGA) utilizing said enzyme.

Process of oxidizing alcohols or carboxylic acids with alcohol/aldehyde

A homogeneous alcohol/aldehyde dehydrogenase isolated from Gluconobacter oxydans DSM 4025 FERM BP-3812 is disclosed. The enzyme is capable in the presence of an electron acceptor, of catalyzing the conversion of L-sorbose to 2-keto-L-gulonic acid via L-sorbosone. The enzyme has a molecular weight of 130,000-140,000 daltons as determined by gel filtration column chromatography, a pyrroloquinoline quinone prosthetic group, an optimum pH for enzyme activity of 7.0 to 9.0, an optimum temperature for enzyme activity from about 20.degree. C. to 40.degree. C. and an isoelectric point of about 4.4. Also disclosed is a process for producing the enzyme and a process for producing aldehydes, carboxylic acids and ketones, especially 2-keto-L-gulonic acid utilizing the enzyme.

Growing community of inventors

Fujisawa, Japan

Akira Asakura