Expression of Mammalian Proteins in Pseudomonas Fluorescens

Abstract

The invention is a process for improved production of a recombinant mammalian protein by expression in a Pseudomonad, particularly in a Pseudomonas fluorescens organism. The process improves production of mammalian proteins, particularly human or human-derived proteins, over known expression systems such as E. coli in comparable circumstances. Processes for improved production of isolated mammalian, particularly human, proteins are provided.

What is claimed is: 
     
         1 . A process for increasing expression of a recombinant mammalian protein comprising:
 a. transforming a  Pseudomonas fluorescens  host cell with a nucleic acid encoding a recombinant mammalian protein; and   b. growing the cell under conditions that allow expression of the recombinant mammalian protein;   wherein the protein is expressed at an increased level when compared to an expression level of the protein under substantially comparable conditions in an  E. coli  expression system.   
     
     
         2 . The process of  claim 1 , further comprising isolating the recombinant mammalian protein. 
     
     
         3 . The process of  claim 2 , further comprising substantially purifying the recombinant mammalian protein. 
     
     
         4 . The process of  claim 1 , wherein the recombinant mammalian protein is present in the host cell in a soluble form. 
     
     
         5 . The process of  claim 1 , wherein the recombinant mammalian protein is present in the cell in an insoluble form. 
     
     
         6 . The process of  claim 1 , wherein the recombinant mammalian protein is present in the cell in an active form. 
     
     
         7 . The process of  claim 1 , wherein the recombinant mammalian protein is a human peptide. 
     
     
         8 . The process of  claim 1  wherein the recombinant protein is produced as more than about 5% total cell protein. 
     
     
         9 . The process of  claim 1  wherein the recombinant protein is produced as more than about 10% total cell protein. 
     
     
         10 . The process of  claim 1  wherein the recombinant protein is produced at a concentration of at least 10 g/L. 
     
     
         11 . The process of  claim 1  wherein the recombinant protein is produced at a concentration of at least 20 g/L. 
     
     
         12 . The process of  claim 1  wherein the recombinant protein is produced at a concentration of at least 40 g/L. 
     
     
         13 . A process for producing a recombinant mammalian protein in a  Pseudomonas fluorescens  host cell comprising:
 a. transforming a host cell with a nucleic acid encoding a recombinant mammalian protein;   b. growing the cell under conditions that allow expression of the recombinant mammalian protein; and   c. isolating the recombinant mammalian protein.   
     
     
         14 . The process of  claim 13 , further comprising substantially purifying the recombinant mammalian protein. 
     
     
         15 . The process of  claim 13 , wherein the recombinant mammalian protein is present in the host cell in a soluble form. 
     
     
         16 . The process of  claim 13 , wherein the recombinant mammalian protein is present in the host cell in an insoluble form. 
     
     
         17 . The process of  claim 13 , wherein the recombinant mammalian protein is present in the host cell in an active form. 
     
     
         18 . The process of  claim 13 , wherein the recombinant mammalian protein is a human peptide. 
     
     
         19 . The process of  claim 13 , wherein the recombinant mammalian protein has a mass of between at least about 1 kD and about 500 kD. 
     
     
         20 . The process of  claim 19 , wherein the recombinant mammalian protein has a mass of greater than about 30 kD. 
     
     
         21 . The process of  claim 13  wherein the recombinant protein is produced as more than about 5% total cell protein. 
     
     
         22 . The process of  claim 13  wherein the recombinant protein is produced as more than about 10% total cell protein. 
     
     
         23 . The process of  claim 13  wherein the recombinant protein is produced at a concentration of at least 10 g/L. 
     
     
         24 . The process of  claim 13  wherein the recombinant protein is produced at a concentration of at least 20g/L. 
     
     
         25 . The process of  claim 13  wherein the recombinant protein is produced at a concentration of at least 40 g/L. 
     
     
         26 . A process for producing a recombinant human protein in a host cell comprising:
 a. transforming a host cell with a nucleic acid encoding a recombinant human peptide; and   b. growing the cell under conditions that allow expression of the recombinant human peptide;   wherein the host cell is  Pseudomonas fluorescens.      
     
     
         27 . The process of  claim 26 , wherein the recombinant human protein is present in soluble form in the host cell. 
     
     
         28 . The process of  claim 26 , wherein the recombinant human protein is present in an active form in the host cell. 
     
     
         29 . A  Pseudomonas fluorescens  cell comprising a nucleic acid encoding a recombinant human peptide. 
     
     
         30 . The cell of  claim 29 , wherein the recombinant human peptide is expressed.