Expression of cold-shock-protein genes from Lactococcus lactis and analysis of the cryoprotection function

Abstract:

[Objective] Bacteria are able to adapt to temperatures far below their optimum growth temperatures, and a set of proteins (cold shock proteins, CSPs) are strongly induced in response to a rapid decrease in growth temperature. We studied the key functions in cryoprotection against damage caused by freezing of these proteins. [Methods] NcoI-HindIII CspC and CspD fragments were cloned respectively between NcoI and HindIII in pNZ8148, the recombinants plasmid were subsequently transformed by electroporation into Lactobacillus lactisNZ9000. Overproduction of CSPs was achieved by the addition of different concentrations of nisin to cultures and was analyzed by SDS-PAGE. In order to study the cryoprotection of CspC and CspD, the growth curves including the control strain and CSP-overproducing strains were developed. The number of colony-forming units was determined just before freezing and after four consecutive freeze-thaw operations. [Results] The 7 kDa cold-shock protein CspD and 6.2 kDa cold-shock protein CspC were identi-fied respectively. [Conclusion] The results indicate that CspC improves the recovery of cells and CspD increases the vi-ability after freezing (30~40 folds).