Designing a Highly Efficient Biosynthetic Route for Lacto-<i>N</i>-Neotetraose Production in <i>Escherichia coli</i>
Pan Zhang, Yingying Zhu, Zeyu Li, Wenli Zhang, Cuie Guang, Wanmeng Mu
Journal of Agricultural and Food Chemistry
Abstract
Recently, the biosynthesis of human milk oligosaccharides (HMOs) has been attracting increasing attention. Lacto-<i>N</i>-neotetraose (LNnT) is one of the most important neutral-core HMOs with promising health effects for infants. It has received Generally Recognized as Safe (GRAS) status and is the second HMO commercially added in infant formula after 2'-fucosyllactose. In previous studies, a series of engineered <i>Escherichia coli</i> strains have been constructed and optimized to produce high titers of precursor lacto-<i>N</i>-triose II. On the basis of these strains, LNnT-producing strains were constructed by overexpressing the β1,4-galactosyltransferase-encoding gene from <i>Aggregatibacter actinomycetemcomitans</i> NUM4039 (Aa<i>-</i>β1,4<i>-</i>GalT). Interestingly, an appreciable LNnT titer was obtained by weakening the metabolic flux of the UDP-GlcNAc pathway and simply overexpressing the essential genes lgtA, galE, and Aa<i>-</i>β1<i>,</i>4<i>-</i>GalT in lacZ-, wecB<i>-</i>, and nagB-deleted <i>E. coli</i>. Subsequently, LNnT synthesis was optimized through balancing the expression of these three biosynthetic enzymes. The optimized strain produced LNnT with an extracellular titer of 12.1 g/L in fed-batch cultivation, with the productivity and specific yield of 0.25 g/L·h and 0.27 g/g dry cell weight, respectively.
Extracted Claims
5 claims extracted from this paper into the knowledge graph
LNnT synthesis optimized through balancing the expression of these three biosynthetic enzymes
“LNnT synthesis was optimized through balancing the expression of these three biosynthetic enzymes.”
Lacto-N-neotetraose (LNnT) received Generally Recognized as Safe (GRAS) status
“It has received Generally Recognized as Safe (GRAS) status and is the second HMO commercially added in infant formula after 2'-fucosyllactose.”
β1,4-galactosyltransferase-encoding gene overexpressed from Aggregatibacter actinomycetemcomitans NUM4039 (Aa-β1,4-GalT)
“LNnT-producing strains were constructed by overexpressing the β1,4-galactosyltransferase-encoding gene from Aggregatibacter actinomycetemcomitans NUM4039 (Aa-β1,4-GalT).”