Metabolic engineering of Selenocysteine Biosynthesis and Insertion Pathway in Lactococcus lactis

Se-enriched lactic acid bacteria (LAB) exist unclear metabolic flow, unstable composition of selenium spectrum and low selenoprotein content such prominent problems caused by complex metabolic pathway and non-specific incorporation of selenium currently. Accordingly, this study reports how to introduce the firstly proposed Selenocysteine Biosynthesis and Insertion Pathway (SBIP) into Lactococcus lactis ( L. lactis ) and specifically guide selenium metabolic flow to direct synthesis of specific selenoprotein with employed multi-level metabolic engineering strategies. In result, the integration of these key factors turned out to facilitate the establishment of SBIP in NZ9000: SelA, SelB, SelC, SelD, GshF and FDH from NZ9000/SBIP up-regulated 8.01, 19.03, 925982.32, 34.51, 31879.16 and 28367.04 multiples compared with NZ9000/p-p; FI/OD ₆₀₀ of NZ9000/SBIP-sfGFP was 362.25 ± 0.43; FDH enzyme activity of NZ9000/SBIP reached 28.11 ± 0.12 mU/mg, and GshF 219.47 ± 0.79 mU/mg under the optimal expression. This first successful implementation of directed synthesis of selenoprotein FDH would indicate a whole new direction to supply Sec-contained proteins through biosynthesis in LAB factory.