초록
To develop an l-lysine high-yielding strain, the enzymes involved in the l-lysine biosynthetic pathway, including aspartokinase III (AK III) and dihydrodipicolinate synthetase (DHDPS) were investigated. Allosteric enzymes involved in l-lysine production from l-lysine producer Escherichia coli LATR11 were sequenced and showed that AK III with mutation T344M or DHDPS with mutation H56K is more conducive to l-lysine production than AK III with mutation M318I or E250K-M318I-G323D or DHDPS with mutation H118Y. Moreover, an l-lysine high-yielding strain was developed from E. coli LATR11 via overexpression of ppc, lysC<SUP>T344M</SUP>, asd, dapA<SUP>H56K</SUP>, dapB, and lysA combined with heterologous expression of Corynebacterium glutamicum ddh. The resulting strain LATR11/pWG-DC<SUP>SM</SUP>A<SUP>SM</SUP>BH<SUB>c.g</SUB>LP showed high l-lysine production (37.2+/-2.3gL<SUP>@?1</SUP>) with productivity (Q<SUB>P</SUB>) of 1.16gL<SUP>@?1</SUP>h<SUP>@?1</SUP> in shake flasks. In fed-batch fermentation, LATR11/pWG-DC<SUP>SM</SUP>A<SUP>SM</SUP>BH<SUB>c.g</SUB>LP produced about 125.6gL<SUP>@?1</SUP> of l-lysine with Q<SUB>P</SUB> of 3.14gL<SUP>@?1</SUP>h<SUP>@?1</SUP> and glucose conversion rate (α) of 58.97% after 40h. From which we got the following conclusions: the enzyme with non-feedback control and high activity, and the high flux through biosynthetic pathway are beneficial to improve l-lysine production in E. coli. These results provide a definite theoretical foundation for breeding amino acid high-yielding strains via genetic engineering from classical producers.