Yeast promoter and use thereof

Abstract

The present invention concerns DNA sequences from the K. lactis PGK gene having transcriptional promoter activity. to expression vectors comprising these sequences and to their use for the production of proteins.

1. An isolated and purified DNA sequence  selected from the group consisting of:
 (a) the sequence  DNA presented in  FIG. 1  (SEQ ID NO: 1);  
 (b) a derivative of the sequence  DNA of (a), wherein the derivative comprises at least nucleotides  1787  to  2242  of SEQ ID NO:  1 ; and  
 (c) a fragment of the sequence  DNA of (a), wherein the fragment comprises at least nucleotides  1787  to  2242  of SEQ ID NO:  1 ;  
 
       wherein said DNA sequence  possesses transcriptional promoter activity. 
     
     
       2. A recombinantly prepared DNA comprising a DNA sequence  according to  claim 1 . 
     
     
       3. A recombinantly prepared DNA according to  claim 2 , further comprising one or more structural genes. 
     
     
       4. A recombinantly prepared DNA according to  claim 3 , wherein said one or more structural genes is human serum albumin. 
     
     
       5. A recombinantly prepared DNA according to  claim 3 , further comprising signals permitting the secretion of the expression product of said structural gene(s). 
     
     
       6. A recombinantly prepared DNA according to  claim 3 , wherein said sequence  DNA is a bidirectional promoter and said genes are  a first structural gene is inserted on each  one side of the promoter and a second structural gene is inserted on the other side of the promoter, in the 2 opposite orientations, provided that said first structural gene and said second structural gene are different. 
     
     
       7. An autonomous or integrative expression plasmid comprising the recombinantly prepared DNA according to  claim 3 . 
     
     
       8. A recombinantly prepared cell containing a DNA sequence  according to  claim 1 . 
     
     
       9. The recombinantly prepared cell according to  claim 8 , characterized in that it is a yeast cell. 
     
     
       10. The recombinantly prepared cell according to  claim 9 , characterized in that said yeast cell is the Kluyveromyces genus. 
     
     
       11. A process for the preparation of a recombinant protein by expression of the  a first gene encoding said protein in a cellular host comprising expression of the said gene under the transcriptional control of a sequence  DNA according to  claim 1 . 
     
     
       12. A process according to  claim 11 , characterized in that the protein is human serum albumin. 
     
     
       13. The process of  claim 11 , further comprising simultaneous expression of isolated and purified genes  a second gene, wherein said sequence  DNA is a bidirectional promoter and said genes are  said first gene is inserted on each  one side of the promoter and said second gene is inserted on the other side of the promoter, in the 2 opposite orientations, provided that said first and second genes are different. 
     
     
       14. An isolated and purified DNA sequence  according to  claim 1 , comprising all or part of  the sequence  DNA presented in FIG.  6 (b) (SEQ ID NO: 2). 
     
     
       15. A recombinantly prepared DNA comprising a DNA sequence  according to  claim 14 . 
     
     
       16. A recombinantly prepared DNA according to  claim 15 , further comprising, one or more structural genes. 
     
     
       17. A recombinantly prepared DNA according to  claim 16 , further comprising signals permitting the secretion of the expression product of the said structural gene(s). 
     
     
       18. An automonous or integrative expression plasmid comprising the recombinantly prepared DNA according to  claim 16 . 
     
     
       19. A recombinantly prepared cell containing a DNA sequence  according to  claim 14 . 
     
     
       20. The recombinantly prepared cell according to  claim 19 , characterized in that it is a yeast cell. 
     
     
       21. The recombinantly prepared cell according to  claim 20 , wherein said yeast cell is of the Kluyveromyces genus. 
     
     
       22. A process for the preparation of a recombinant protein comprising expression of the  a first gene encoding said protein in a cellular host wherein the expression of said gene is under the transcriptional control of a sequence  DNA according to  claim 14 . 
     
     
       23. The process of  claim 22 , further comprising simultaneous expression of isolated and purified genes  a second gene, wherein said sequence  DNA is a bidirectional promoter and said genes are  said first gene is inserted on each  one side of the promoter and said second gene is inserted on the other side of the promoter, in the 2 opposite orientations, provided that said first and second genes are different.