Modified Kluyveromyces yeasts, their preparation and use

Abstract

The present invention concerns yeasts of the genus Kluyveromyces having one or more genetic modifications of at least one gene coding for a protease, said gene reducing or modifying the proteolytic actively of said yeasts, as well as their use as a cellular host for the secretion of recombinant proteins.

We claim:  
     
       1. A Kluyverornyces  Kluyveromyces yeast comprising a genetic modification in its PRA1, PRB1 or PRC1 gini  gene, wherein said modification results in loss of protease activity encoded by said gene. 
     
     
       2. The Kluyverormyces yeast according to claim  1 , wherein the modification is in 
       a) protein coding region of said gene;  
       b) a region responsible for transcriptional regulation of said gene; or  
       c) both a protein coding region of said gene and a region responsible for transcriptional regulation of said gene.  
     
     
       3. The Kluyveromyces yeast according to claim  1 , selected from the group consisting of K. lactis, K. fragilis, K, drosophilarum, and K. waltii. 
     
     
       4. A process for preparing the Kluyveromyces yeast of claim  1  comprising replacing all or part of said gene with a modified version thereof. 
     
     
       5. A Kluyveromyces yeast comprising a genetic modification in its PRA1, PRB1, or PRC1 gene, wherein said modification results in loss of protease activity encoded by said gene, and an exogenous DNA sequence comprising a nucleic acid sequence encoding at least one protein. 
     
     
       6. The Kluyveromyces yeast according to claim  5 , wherein the exogenous DNA sequence further comprises a region for initiation of transcription and translation operably linked to the nucleic acid sequence encoding the at least one protein. 
     
     
       7. The Kluyveromyces yeast according to claim  5 , wherein the exogenous sequence comprises a region encoding an exporting sequence for secretion of the protein. 
     
     
       8. The Kluyveromyces yeast according to claim  5 , wherein the exogenous DNA sequence is part of an autonomously replicating vector or is integrated into a chromosome. 
     
     
       9. A process for producing recombinant proteins comprising culturing the Kluyverornyces  Kluyveromyces yeast of claim  5  under conditions for expression of said exogenous DNA sequence. 
     
     
       10. A process for producing recombinant proteins comprising culturing the Kluyveromyces yeast of claim  6  under conditions for expression of said exogenous DNA sequence. 
     
     
       11. A process for producing recombinant proteins comprising culturing the Kluyverormyces yeast of claim  7  under conditions for expression of said exogenous DNA sequence. 
     
     
       12. The Kluyveromyces yeast according to claim  5 , wherein the exogenous DNA sequence encodes a protein of pharmaceutical interest. 
     
     
       13. A process for producing recombinant proteins comprising culturing the Kluyveromyces yeast of claim  12  under conditions for expression of said exogenous DNA sequence.