Patent EA-003773-B1 (EA)
Title: Dna sequences for 5s clavam biosynthesis in streptomyces clavuligerus microorganisms, streptomyces clavuligerus microorganism strains, containing said dna sequences and a process for improving 5r clavam production, in particular, clavulanic acid in streptomyces clavuligerus microorganisms
Inventor: BARTON BARRY
IPC: A61P31/04
Filed: 1998-02-02
Abstract: 1. A DNA molecule comprising a cas1 gene and flanking regions specific for 5S clavam biosynthesis in Streptomyces clavuligerus microorganisms and which is not essential for 5R clavam biosynthesis having nucleid sequence (SEQ ID No. 1), identified in Figure 1 or a DNA molecule which is hybridised under conditions of high stringency with said DHA molecule. 2. The DNA molecule which is an open reading frame specific for 5S clavam biosynthesis in Streptomyces clavuligerus microorganisms and which is not essential for 5R clavam biosynthesis selected from the group consisting of: a). orfup1, having the sequence (SEQ ID No. 4); b). orfup2, having the sequence (SEQ ID No. 3); c). orfup3, having the sequence (SEQ ID No. 2); d). orfdwn1, having the sequence (SEQ ID No. 5); e). orfdwn2, having the sequence (SEQ ID No. 6); f). orfdwn3, having the sequence (SEQ ID No. 7). 3. A vector comprising the DNA molecule of any one of claims 1 or 2. 4. Plasmid comprising one or more genes specific for 5S clavam biosynthesis from which one or more genes has been disrupted or otherwise made defective wherein the genes correspond to open reading frame selected from the group consisting of: a). orfup1, having the sequence (SEQ ID No. 4); b). orfup2, having the sequence (SEQ ID No. 3); c). orfup3, having the sequence (SEQ ID No. 2); d). orfdwn1, having the sequence (SEQ ID No. 5); e). orfdwn2, having the sequence (SEQ ID No. 6); f). orfdwn3, having the sequence (SEQ ID No. 7). 5. The plasmid of claim 4 which is pCEC060, pCEC061, pCEC056, pCEC057 or pDES3. 6. Streptomyces clavuligerus microorganism strain comprising the plasmid of claim 4. 7. Streptomyces clavuligerus microorganism strain comprising one or more genes specific for 5S clavam biosynthesis from which one or more genes has been disrupted or otherwise made defective wherein the genes have open reading frame selected from the group consisting of: a). orfup1, having the sequence (SEQ ID No. 4); b). orfup2, having the sequence (SEQ ID No. 3); c). orfup3, having the sequence (SEQ ID No. 2); d). orfdwn1, having the sequence (SEQ ID No. 5); e). orfdwn2, having the sequence (SEQ ID No. 6); f). orfdwn3, having the sequence (SEQ ID No. 7); wherein said disrupted or otherwise made defective genes have been included in said Streptomyces clavuligerus microorganism strain by changing genes using the plasmid of claim 4. 8. A process for improving 5R clavam production in a Streptomyces clavuligerus microorganism strain comprising the steps of: a). disrupting or otherwise making defective DNA of claim 1 or 2; b). introducing disrupted or defective DNA obtained after a) in Streptomyces clavuligerus microorganism strain capable of producing 5R clavam; and c). fermentation of Streptomyces clavuligerus microorganism strain obtained after b) under conditions suitable for producing 5R clavam. 9. The process of claim 8, wherein said DNA corresponds to open reading frames ortup3, ortup2, orfupl, orfdwn 1, orfdwn2 or orfdwn3. 10. The process of claim 8 or 9, wherein the 5R clavam is clavulanic acid.