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KMID : 1094720170220040390
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Biotechnology and Bioprocess Engineering
2017 Volume.22 No. 4 p.390 ~ p.396
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glyA gene knock-out in Escherichia coli enhances L-serine production without glycine addition
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Zhang Ya
Kang Pei
Liu Shuang
Zhao Yujiao
Wang Zhiwen
Chen Tao
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Abstract
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In E. coli, glyA encodes for serine hydroxymethyltransferase (SHMT), which converts L-serine to glycine. When engineering L-serine-producing strains, it is therefore favorable to inactivate glyA to prevent L-serine degradation. However, most glyA knockout strains exhibit slow cell growth because of the resulting lack of glycine and C1 units. To overcome this problem, we overexpressed the gcvTHP genes of the glycine cleavage system (GCV), to increase the C1 supply before glyA was knocked out. Subsequently, the kbl and tdh genes were overexpressed to provide additional glycine via the L-threonine degradation pathway, thus restoring normal cell growth independent of glycine addition. Finally, the plasmid pPK10 was introduced to overexpress pgk, serA Δ197 , serC and serB, and the resulting strain E4G2 (pPK10) accumulated 266.3 mg/L of L-serine in a semi-defined medium without adding glycine, which was 3.18-fold higher than the production achieved by the control strain E3 (pPK10). This strategy can accordingly be applied to disrupt the L-serine degradation pathway in industrial production strains without causing negative side-effects, ultimately making L-serine production more efficient.
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KEYWORD
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Escherichia coli, glyA, glycine cleavage system, kbl-tdh, L-serine, metabolic engineering
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