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Applied and Environmental Microbiology, September 2000, p. 3756-3763, Vol. 66, No. 9
0099-2240/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Physiological and Regulatory Effects of Controlled Overproduction of Five Cold Shock Proteins of Lactococcus lactis MG1363

Jeroen A. Wouters,1,2 Marielle Mailhes,1 Frank M. Rombouts,1 Willem M. de Vos,2 Oscar P. Kuipers,2,dagger and Tjakko Abee1,*

Laboratory of Food Microbiology, Wageningen University and Research Center, Wageningen,1 and Microbial Ingredients Section, NIZO Food Research, Ede,2 The Netherlands

Received 18 January 2000/Accepted 13 June 2000

The physiological and regulatory effects of overproduction of five cold shock proteins (CSPs) of Lactococcus lactis were studied. CspB, CspD, and CspE could be overproduced at high levels (up to 19% of the total protein), whereas for CspA and CspC limited overproduction (0.3 to 0.5% of the total protein) was obtained. Northern blot analysis revealed low abundance of the cspC transcript, indicating that the stability of cspC mRNA is low. The limited overproduction of CspA is likely to be caused by low stability of CspA since when there was an Arg-Pro mutation at position 58, the level of CspA production increased. Using two-dimensional gel electrophoresis, it was found that upon overproduction of the CSPs several proteins, including a number of cold-induced proteins of L. lactis, were induced. Strikingly, upon overproduction of CspC induction of CspB, putative CspF, and putative CspG was also observed. Overproduction of CspB and overproduction of CspE result in increased survival when L. lactis is frozen (maximum increases, 10- and 5-fold, respectively, after 4 freeze-thaw cycles). It is concluded that in L. lactis CSPs play a regulatory role in the cascade of events that are initiated by cold shock treatment and that they either have a direct protective effect during freezing (e.g., RNA stabilization) or induce other factors involved in the freeze-adaptive response or both.

* Corresponding author. Mailing address: Laboratory of Food Microbiology, Wageningen University and Research Center, Bomenweg 2, 6703 HD Wageningen, The Netherlands. Phone: 31-317-484981. Fax: 31-317-484893. E-mail: Tjakko.Abee{at}micro.fdsci.wau.nl.

dagger Present address: Department of Genetics, University of Groningen, 9750 AA Haren, The Netherlands.

Applied and Environmental Microbiology, September 2000, p. 3756-3763, Vol. 66, No. 9
0099-2240/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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