Patent ID: 8916358

Claim:
A method of producing a product of a biosynthetic pathway of interest, the method comprising: (i) growing genetically modified bacterial cells that comprise nucleic acids that encode: (a) enzymes of the biosynthetic pathway of interest that catalyze production of the product, (b) a targeted enzyme that competes for substrates or cofactors with an enzyme in the biosynthetic pathway of interest, decreases the overall yield and/or the rate of production of the product, and includes a site-specific protease recognition sequence, and (c) a site-specific protease that cleaves the site-specific protease recognition sequence of the targeted enzyme and is genetically modified to include a periplasmic targeting sequence; (ii) lysing at least a portion of the bacterial cells to produce a lysate, thereby bringing the site-specific protease into contact with the targeted enzyme to cleave and inactivate or reduce the activity of the targeted enzyme; (iii) adding substrate to the lysate; and (iv) incubating the lysate and substrate, thereby producing the product of the biosynthetic pathway of interest.