ENHANCED PRODUCTION OF L-AMINOADIPIC ACID IN Escherichia coli BY METABOLIC ENGINEERING OF LYSINE AND ADIPIC ACID BIOSYNTHESIS

Abstract

L- Aminoadipic acid (L- AAA), a non- protein structure amino acid, is an important intermediate for many medicinal compounds such as antirheumatic drug, psoriasis and carcinostatic drug as well as a precursor in the production of β-lactam antibiotics. L-Lysine is known as one of precursors for L-AAA synthesis in microorganisms. To increase L-AAA production in Escherichia coli, releasing of allosteric inhibition of the enzymes in L-lysine biosynthesis pathway should be performed. pD-Y*D*LP containing V339A mutated lysC and E84T mutated dapA genes encoding for L-lysine feedback resistant aspartokinase III and dihydrodipicolinate synthase from E. coli, respectively, along with L–AAA synthesis genes, lysdh which encodes lysine dehydrogenase from Acromobacter denitrifican and pcd which encodes piperideine-6-carboxylate dehydrogenase from Pseudomonas putida was constructed. Proteins expression of aspartokinase III and dihydrodipicolinate synthase as well as the activity of aspartokinase III from the recombinant clone were not clearly detected. However, HPLC and TLC analysis indicated that the presence of V399A lysC and E84T dapA under T7 promoter of pRSF-Duet1 could elevate L-AAA titer when the recombinant clone was cultured in minimal medium.