Method for producing soluble recombinant interferon protein without denaturing

摘要

The present invention relates to the field of recombinant protein production in bacterial hosts. It further relates to extraction of soluble, active recombinant protein from an insoluble fraction without the use of denaturation and without the need for a refolding step. In particular, the present invention relates to a production process for obtaining high levels a soluble recombinant Type 1 interferon protein from a bacterial host.

主权项

1. A method for producing a recombinant Type 1 interferon protein, said method comprising:
expressing the recombinant Type I interferon protein by culturing a Pseudomonas or E. coli host cell containing an expression construct comprising a nucleotide sequence encoding the recombinant Type I interferon protein that has been optimized for expression in the host cell, wherein the recombinant Type I interferon protein is expressed in the cytoplasm of the host cell; lysing the cultured host cell; obtaining an insoluble fraction and a soluble fraction from the lysed host cell; extracting the insoluble fraction by subjecting it to non-denaturing extraction conditions; wherein the non-denaturing extraction conditions comprise a Zwitterionic detergent at a concentration of about 0.5% to about 2%, to obtain soluble recombinant Type 1 interferon protein without the use of a renaturing or refolding step; separating the obtained soluble recombinant Type 1 interferon protein from other protein and cellular debris; and measuring the activity of the separated soluble recombinant Type 1 interferon protein, wherein about 80% to about 100% of the separated soluble recombinant Type 1 interferon protein is determined to be active.