BACILLUS HOSTS FOR PHARMACEUTICAL PROTEIN SECRETION

BACILLUS SUBTILIS IS AN ADVANTAGEOUS HOST ORGANISM FOR THE PRODUCTION OF FOREIGN PROTEINS AT LOW COST BECAUSE IT HAS THE CAPACITY TO SECRETE HIGH LEVELS OF PROTEINS INTO THE GROWTH MEDIUM. SEVERAL PROTEINS OF BACTERIAL ORIGIN ENCODEDBY PLASMID-BORNE RECOMBINANT GENES HAVE BEEN SECRETED BY B. SUBTILIS STRAINS, AND B. SUBTILIS CAN SECRETE SEVERAL PROTEINS OF EUKARYOTIC ORIGIN, ALTHOUGH THIS HAS NOT BEEN AS SUCCESSFUL AS WITH PROTEINS OF BACTERIAL ORIGIN. THIS PROPOSAL IS DESIGNED TO EXAMINE THE HYPOTHESIS THAT THE DISCRIMINATION AGAINST PROTEINS OF EUKARYOTIC ORIGIN IN B. SUBTILIS IS RELATED TO THE TIMING OF PROTEOLYTIC REMOVAL OF THE N-TERMINAL SECRETION SIGNAL PEPTIDE, WHICH OCCURS LATER,RELATIVE TO THE SYNTHESIS OF THE PROTEIN, IN BACTERIA THAN IN EUKARYOTIC SYSTEMS. OVERPRODUCTION OF THE SIGNAL PEPTIDASE ENZYME IN B. SUBTILIS IS PROPOSED, IN ORDER TO ACCELERATE SIGNAL REMOVAL FROM SECRETED PROTEINS. TO ACCOMPLISH THIS, CLONING OF THE B. SUBTILIS GENE ENCODING SIGNAL PEPTIDASE IS PROPOSED. THE LONG-TERM OBJECTIVE OF THIS PROPOSAL IS TO CONSTRUCT B. SUBTILIS HOST STRAINS THAT CAN SECRETE PROTEINS OF EUKARYOTIC ORIGIN AT LEVELS COMPARABLE TO THOSE ACHIEVED FOR PROTEINS OF BACTERIAL ORIGIN. SUCH STRAINS WOULD MAKE POSSIBLE THE PRODUCTION AT LOW COST OF PROTEINS OF PHARMACEUTICAL INTEREST, SOME OF WHICH CANNOT BE PRODUCED IN ACTIVE FORM IN CURRENTLY AVAILABLE MICROBIAL SYSTEMS.