Construction and application of a food-grade expression system for Lactococcus lactis

Area of Science:

• Microbiology
• Biotechnology
• Food Science

Background:

• Lactococcus lactis is a widely used food-grade bacterium for industrial applications.
• Development of efficient host/vector systems is crucial for its biotechnological potential.
• Alanine racemase gene (alr) can serve as a valuable complementation marker for bacterial genetics.

Purpose of the Study:

• To construct a novel food-grade host/vector expression system for Lactococcus lactis.
• To utilize the alanine racemase gene (alr) as a selectable marker for system development.
• To demonstrate the system's efficacy in overexpressing foreign proteins.

Main Methods:

• Construction of an alanine racemase auxotrophic mutant (L. lactis NZ9000Δalr) using double-crossover recombination.
• Development of a food-grade vector (pALR) containing lactococcal DNA elements and the alr gene.
• Overexpression of green fluorescent protein and porcine circovirus type II capsid protein using the nisin-inducible promoter PnisA.

Main Results:

• Successfully generated the L. lactis NZ9000Δalr auxotrophic mutant.
• Developed a functional food-grade expression vector (pALR) with lactococcal DNA elements.
• Achieved efficient overexpression of target proteins (GFP and PCV2 capsid protein) under nisin induction.

Conclusions:

• The developed food-grade host/vector system for Lactococcus lactis is effective for protein expression.
• This system holds significant potential as an antigen delivery vehicle.
• The system is suitable for manufacturing ingredients for the food industry.