DESIGN OF Escherichia coli COCULTURE SYSTEM TO PRODUCE MULTI-METHYL-BRANCHED ESTERS

GRAMAJO, HUGO; BRACALENTE, FERNANDO; ARABOLAZA, ANA; SABATINI, MARTÍN

Paraná

Congreso; LIV Reunión Anual Sociedad Argentina de Investigación en Bioquímica y Biología Molecula; 2018

Sociedad Argentina de Investigación Bioquímica y Biología Molecular (SAIB)

Microbial lipid production represents a potential alternative feedstock for oleochemical industries. In a previous work, we engineered into E. coli a polyketide synthase (PKS)-based biosynthetic pathway from Mycobacterium tuberculosis and redefined its biological role towards the production of a variety of multi-methyl-branched esters (MBE).To produce the MBE, it was necessary to supplement the culture media with the alcohol that esterifies to the multi-methyl-fatty acid (MBFA) synthesized by the PKS. With the aim to develop a whole de novo bioprocess (i.e. without usingexogenous alcohol), an E. coli co-culture system was engineered to modularize the ?alcohol biosynthetic pathway? and the ?PKS-based biosynthetic pathway?. Thus, the MBE producing strain (RQ5 pMB22) was cultivated with an isobutanolproducing E. coli strain. To achieve this goal, we first generated this new strain, CB1, by integrating into the chromosome of an E. coli BL21 strain, the genes encoding the enzymes necessary for the biosynthesis of isobutanol from glucose.CB1 produced 18,75 ± 0,45 mM of isobutanol after 24h of induction, and when it was cocultivated with RQ5 pMB22 in a minimal M9 medium, the consortium RQ5/CB1, at the best inoculation ratio of 1:0.5, produced 30.48 ± 2.02 μg MBE ml-1.This yield was comparable to that exhibited by a monoculture based on the single-cell RQ5 pMB22 with the external addition of isobutanol into the growth media.