Production of bovine prochymosin in Lactococcus lactis

Abstract:

Abstract:[Objective]Bovine prochymosin is expressed by the nisin controlled gene expression system in Lactococcus lactis.[Methods]we amplified bovine prochymosin gene from pS19-PPC vector by PCR,and ligated the gene with expression vector pNZ8148.Ligated products were electrotransformated into Lactococcus lactis NZ9000.Then we identified transformants by restriction,PCR and sequencing, and molecular weight and immune characteristics of expression product by SDS-PAGE and Western blot after inducting using nisin.Eventually we tested milk-clotting activity after purification.[Results]Recombinant bovine prochymosin was not significant difference in molecular weight,immune characteristics, bioactivity and sensitivity of inhibitors comparison to nature bovine prochymosin.The milk-clotting activity of recombinant bovine prochymosin was 2×103 international milk-clotting unit per millilitre.[Conclusion]We expressed recombinant bovine prochymosin of milk-clotting activity in Lactococcus lactis. This study suggested potential application in cheese manufacture as a new method,which could regard Lactococcus lactis as starter of milk and host of expression bovine prochymosin.

Key words:Keyword: Bovine prochymosin; Lactococcus lactis; Western blot; milk-clotting activity