Yeast host, transformant and method for producing heterologous proteins

Abstract

A method of constructing a host for expression of an exogenous gene which comprises deleting or inactivating at least one gene selected from the protease-associated genes of Schizosaccharomyces pombe such as psp3 (SPAC1006.01), sxa2 (SPAC1296.03c), ppp51 (SPAC22G7.01c) or ppp52 (SPBC18A7.01). An isolated or purified host cell in which a protease-associated gene is deleted or inactivated, a transformant obtained by introducing an exogenous gene into the host cell and a method of producing an exogenous or heterologous protein using the transformant.

1 . A method of constructing a  Schizosaccharomyces pombe  host cell for expression of a recombinantly introduced foreign gene, comprising:
 deleting or inactivating at least one target gene selected from the groups consisting of genes encoding serine proteases (serine protease gene family), genes encoding amino peptidases (amino peptidase gene family), genes encoding carboxy peptidases (carboxy peptidase gene family) and genes encoding dipeptidases (dipeptidase gene family).   
     
     
         2 . The method according to  claim 1 , wherein the target gene is at least one gene selected from the group consisting of psp3 (SPAC1006.01), sxa2 (SPAC1296.03c), ppp51 (SPAC22G7.01c) and ppp52 (SPBC18A7.01). 
     
     
         3 .- 5 . (canceled) 
     
     
         6 . The method according to  claim 1 , wherein the genes are deleted or inactivated by replacing their open reading frames (ORFs) with marker genes. 
     
     
         7 . A  Schizosaccharomyces pombe  host for expression of a recombinantly introduced foreign gene, in which at least one gene selected from the group consisting of psp3 (SPAC1006.01), sxa2 (SPAC1296.03c), ppp51 (SPAC22G7.01c) and ppp52 (SPBC18A7.01) is deleted or inactivated. 
     
     
         8 .- 10 . (canceled) 
     
     
         11 . A transformant obtained by introducing a gene encoding a heterologous protein into the host as defined above in  claim 7 . 
     
     
         12 . The transformant according to  claim 11 , wherein a secretion signal gene is introduced with the gene encoding a heterologous protein. 
     
     
         13 . A method of producing a heterologous protein, which comprises culturing the transformant as defined in  claim 11  to allow it to produce the heterologous protein, and recovering the heterologous protein. 
     
     
         14 . A method of producing a heterologous protein, which comprises culturing the transformant as defined in  claim 12  to allow it to produce the heterologous protein and secret the heterologous protein in the culture, and recovering the heterologous protein from the culture. 
     
     
         15 . The method according to  claim 13 , wherein according to 13 mentioned above, wherein the heterologous protein is human growth hormone (hGH). 
     
     
         16 . An isolated or purified  Schizosaccharomyces pombe  cell produced by deleting or inactivating from a corresponding  Schizosaccharomyces pombe  parent strain at least one gene selected from the group consisting of:
 a metalloprotease family gene selected from cdb4 (SPAC23H4.09), mast (SPBC18E5.12c), pgp1 (SPCC1259.10), ppp20 (SPAC4F10.02), ppp22 (SPBC14C8.03), ppp51 (SPC22G7.01c), ppp52 (SPBC18A7.01) and ppp53 (SPAP14E8.04);   a serine protease family gene selected from isp6 (SPAC4A8.04), ppp16 (SPBC1711.12), psp3 (SPAC1006.01) and sxa2 (SPAC1296.03c);   a cysteine protease family gene selected from ppp80 (SPAC19B12.08), pca1 (SPCC1840.04), cut1 (SPCC5E4.04) and gpi8 (SPCC11E10.02c); and   a aspartyl protease family gene selected from sxa1 (SPAC26A3.01), yps1 (SPCC1795.09) and ppp81 (SPAC25B8.17).   
     
     
         17 . The isolated or purified  Schizosaccharomyces pombe  cell of  claim 16  produced by deleting or inactivating at least one gene selected from the group consisting of psp3 (SPAC1006.01), sxa2 (SPAC1296.03c), ppp51 (SPAC22G7.01c) and ppp52 (SPBC18A7.01) from a corresponding  Schizosaccharomyces pombe  parent strain. 
     
     
         18 . The isolated or purified  Schizosaccharomyces pombe  cell of  claim 16 , wherein said at least one gene is deleted. 
     
     
         19 . The isolated or purified  Schizosaccharomyces pombe  cell of  claim 16 , wherein said at least one gene is inactivated. 
     
     
         20 . The isolated or purified  Schizosaccharomyces pombe  cell of  claim 16 , wherein said at least one gene comprises psp3 (SPAC1006.01). 
     
     
         21 . The isolated or purified  Schizosaccharomyces pombe  cell of  claim 16 , wherein said at least one gene consists of psp3 (SPAC1006.01). 
     
     
         22 . The isolated or purified  Schizosaccharomyces pombe  cell of  claim 16 , wherein said at least one gene comprises sxa2 (SPAC1296.03c). 
     
     
         23 . The isolated or purified  Schizosaccharomyces pombe  cell of  claim 16 , wherein said at least one gene consists of sxa2 (SPAC1296.03c). 
     
     
         24 . The isolated or purified  Schizosaccharomyces pombe  cell of  claim 16 , wherein said at least one gene comprise ppp51 (SPAC22G7.01c). 
     
     
         25 . The isolated or purified  Schizosaccharomyces pombe  cell of  claim 16 , wherein said at least one gene consists of ppp51 (SPAC22G7.01c). 
     
     
         26 . The isolated or purified  Schizosaccharomyces pombe  cell of  claim 16 , wherein said at least one gene comprises ppp52 (SPBC18A7.01). 
     
     
         27 . The isolated or purified  Schizosaccharomyces pombe  cell of  claim 16 , wherein said at least one gene consists of ppp52 (SPBC18A7.01). 
     
     
         28 . The isolated or purified  Schizosaccharomyces pombe  cell of  claim 16 , further comprising a polynucleotide sequence or vector encoding an exogenous polypeptide. 
     
     
         29 . A method for making an exogenous polypeptide comprising
 transforming the isolated or purified  Schizosaccharomyces pombe  cell of  claim 16  with a polynucleotide or vector encoding said exogenous proteins under conditions suitable for expression of said exogenous polypeptide and   recovering said exogenous polypeptide.