Skip to content
1887

Microbiology Society logo Microbiology

Volume 145, Issue 11

Analysis of the role of 7 kDa cold-shock proteins of MG1363 in cryoprotection Free

Abstract

Low-temperature adaptation and cryoprotection were studied in the lactic acid bacterium MG1363. An approximately 100-fold increased survival after freezing was observed when cells were shocked to 10 °C for 4 h compared to mid-exponential-phase cells grown at 30 °C, indicating an active protection against freezing. Using two-dimensional gel electrophoresis a group of 7 kDa cold-induced proteins (CSPs) was identified that corresponds to a previously described family of genes of MG1363 (Wouters , 1998 , 144, 2885–2893). The 7 kDa CSPs appeared to be the most strongly induced proteins upon cold shock to 10 °C. Northern blotting and two-dimensional gel electrophoresis showed that the genes were maximally expressed at 10 °C, while induction was lower at 20 and 4 °C. However, pre-incubation at 20 and 4 °C, as well as stationary-phase conditions, also induced cryoprotection (approx. 30-, 130- and 20-fold, respectively, compared to 30 °C mid-exponential phase). For all treatments leading to an increased freeze survival (exposure to 4, 10 and 20 °C and stationary-phase conditions), increased levels of three proteins (26, 43 and 45 kDa) were observed for which a role in cryoprotection might be suggested. Increased freeze survival coincides with increased CSP expression, except for stationary-phase conditions. However, the level of observed freeze protection does not directly correlate with the gene expression levels. In addition, for the first time specific overproduction of a CSP in relation to freeze survival was studied. This revealed that cells overproducing CspD at 30 °C show a 2–10-fold increased survival after freezing compared to control cells. This indicates that the 7 kDa cold-shock protein CspD may enhance the survival capacity after freezing but that other factors supply additional cryoprotection.

  • Received:
  • Accepted:
  • Revised:
  • Published Online:
Keyword(s): 2D-EF, two-dimensional gel electrophoresis , cold-shock proteins , cryoprotection , CSP, cold-shock protein , LAB, lactic acid bacteria , Lactococcus lactis and low-temperature adaptation
© Microbiology Society
Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-145-11-3185
1999-11-01
2025-01-18
Loading full text...

Full text loading...

/deliver/fulltext/micro/145/11/1453185a.html?itemId=/content/journal/micro/10.1099/00221287-145-11-3185&mimeType=html&fmt=ahah

References

  1. Becker L. A., Sevket Cetin M., Hutkins R. W., Benson A. K. 1998; Identification of the gene encoding the alternative sigma factor σB from Listeria monocytogenes and its role in osmotolerance. J Bacteriol 180:4547–4554
     [Google Scholar]
  2. Blum H., Beier H., Gross H. J. 1987; Improved silver staining of plant proteins, RNA and DNA in polyacrylamide gels. Electrophoresis 8:93–99 [CrossRef]
     [Google Scholar]
  3. El-kest S. E., Marth E. H. 1992; Freezing of Listeria monocytogenes and other microorganisms: a review. J Food Protein 55:639–648
     [Google Scholar]
  4. Etchegaray J.-P., Inouye M. 1999; CspA, CspB, and CspG, major cold shock proteins of Escherichia coli, are induced at low temperature under conditions that completely block protein synthesis. J Bacteriol 181:1827–1830
     [Google Scholar]
  5. Franks F. 1995; Protein destabilization at low temperatures. Adv Protein Chem 46:105–139
     [Google Scholar]
  6. Gansel X., Hartke A., Boutibonnes P., Auffray Y. 1993; Nucleotide sequence of the Lactococcus lactis NCDO763 (ML3) rpoD gene. Biochim Biophys Acta 1216:115–118 [CrossRef]
     [Google Scholar]
  7. Gasson M. J. 1983; Plasmid complements of Streptococcus lactis NCDO712 and other lactic streptococci after protoplast-induced curing. J Bacteriol 154:1–9
     [Google Scholar]
  8. Goldstein J., Politt N. S., Inouye M. 1990; Major cold shock protein of Escherichia coli. Proc Natl Acad Sci USA 87:283–287 [CrossRef]
     [Google Scholar]
  9. Graumann P., Marahiel M. A. 1996; Some like it cold: response of microorganisms to cold shock. Arch Microbiol 166:293–300 [CrossRef]
     [Google Scholar]
  10. Graumann P., Schröder K., Schmid R., Marahiel M. A. 1996; Cold shock stress-induced proteins in Bacillus subtilis. J Bacteriol 178:4611–4619
     [Google Scholar]
  11. Graumann P., Wendrich T. M., Weber M. H. W., Schröder K., Marahiel M. A. 1997; A family of cold shock proteins in Bacillus subtilis is essential for cellular growth and for efficient protein synthesis at optimal and low temperatures. Mol Microbiol 25:741–756 [CrossRef]
     [Google Scholar]
  12. Hartke A., Bouche S., Gansel X., Boutibonnes P., Auffray Y. 1994; Starvation-induced stress resistance in Lactococcus lactis subsp. lactis IL1403. Appl Environ Microbiol 60:3474–3478
     [Google Scholar]
  13. Hecker M., Schumann W., Völker U. 1996; Heat-shock and general stress response in Bacillus subtilis. Mol Microbiol 19:417–428 [CrossRef]
     [Google Scholar]
  14. Jiang W., Hou Y., Inouye M. 1997; CspA, the major cold-shock protein of Escherichia coli, is an mRNA chaperone. J Biol Chem 272:196–202 [CrossRef]
     [Google Scholar]
  15. Jones P. G., Inouye M. 1994; The cold-shock response – a hot topic. Mol Microbiol 11:811–818 [CrossRef]
     [Google Scholar]
  16. Jones P. G., Krah R., Tafuri S. R., Wolffe A. P. 1992; DNA gyrase, CS7·4, and the cold shock response in Escherichia coli. . J Bacteriol 174:5798–5802
     [Google Scholar]
  17. Kim W. S., Dunn N. W. 1997; Identification of a cold shock gene in lactic acid bacteria and the effect of cold shock on cryotolerance. Curr Microbiol 35:59–63 [CrossRef]
     [Google Scholar]
  18. Kolter R., Stiegele D. A., Tormo A. 1993; The stationary phase of the bacterial life-cycle. Annu Rev Microbiol 47:855–874 [CrossRef]
     [Google Scholar]
  19. Kuipers O. P., Beerthuyzen M. M., Siezen R. J., de Vos W. M. 1993; Characterization of the nisin gene cluster nisABTCIPR of Lactococcus lactis: requirement of expression of the nisA and nisI genes for development of immunity. Eur J Biochem 216:281–291 [CrossRef]
     [Google Scholar]
  20. Kuipers O. P., Beerthuyzen M. M., de Ruyter P. G. G. A., Luesink E. J., de Vos W. M. 1995; Autoregulation of nisin biosynthesis in Lactococcus lactis by signal transduction. J Biol Chem 270:27299–27304 [CrossRef]
     [Google Scholar]
  21. LaTeana A., Brandi A., Falconi M., Spurio R., Pon C. L., Gualerzi C. O. 1991; Identification of a cold shock transcriptional enhancer of the Escherichia coli major cold shock gene encoding nucleoid protein H-NS. Proc Natl Acad Sci USA 88:10907–10911 [CrossRef]
     [Google Scholar]
  22. Lee S. J., Xie A., Jiang W., Etchegaray J., Jones P. G., Inouye M. 1994; Family of the major cold-shock protein, CspA (CS7.4), of Escherichia coli, whose members show a high sequence similarity with the eukaryotic Y-box binding proteins. Mol Microbiol 11:833–839 [CrossRef]
     [Google Scholar]
  23. Nakashima K., Kanamaru K., Mizuno T., Horikoshi K. 1996; A novel member of the cspA family of genes that is induced by cold shock in Escherichia coli. . J Bacteriol 178:2994–2997
     [Google Scholar]
  24. O’Farrell P. H. 1975; High resolution two-dimensional electrophoresis of proteins. J Biol Chem 250:4007–4021
     [Google Scholar]
  25. Panoff J.-M., Thammavongs B., Laplace J.-M., Hartke A., Boutibonnes P., Auffray Y. 1995; Cryotolerance and cold adaptation in Lactococcus lactis subsp. lactis IL1403. Cryobiology 32:516–520 [CrossRef]
     [Google Scholar]
  26. Rallu F., Gruss A., Maguin E. 1996; Lactococcus lactis and stress. Antonie Leeuwenhoek 70:243–251 [CrossRef]
     [Google Scholar]
  27. van Rooijen R. J., de Vos W. M. 1990; Molecular cloning, transcriptional analysis, and nucleotide sequence of lacR, a gene encoding the repressor of the lactose phosphotransferase system of Lactococcus lactis. . J Biol Chem 265:18499–18503
     [Google Scholar]
  28. de Ruyter P. G. G. A., Kuipers O. P., de Vos W. M. 1996; Controlled gene expression systems for Lactococcus lactis with the food-grade inducer nisin. Appl Environ Microbiol 62:3662–3667
     [Google Scholar]
  29. Schägger H., von Jagow G. 1987; Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis for the separation of proteins in the range from 1 to 100 kDa. Anal Biochem 166:368–379 [CrossRef]
     [Google Scholar]
  30. Thammavongs B., Corroler D., Panoff J.-M., Auffray Y., Boutibonnes P. 1996; Physiological response of Enterococcus faecalis JH2-2 to cold shock: growth at low temperatures and freezing/thawing challenge. Lett Appl Microbiol 23:398–402 [CrossRef]
     [Google Scholar]
  31. Willimsky G., Bang H., Fischer G., Marahiel M. A. 1992; Characterization of cspB, a Bacillus subtilis inducible cold shock gene affecting cell viability at low temperatures. J Bacteriol 174:6326–6335
     [Google Scholar]
  32. Wouters J. A., Sanders J.-W., Kok J., de Vos W. M., Kuipers O. P., Abee T. 1998; Clustered organization and transcriptional analysis of a family of five csp genes of Lactococcus lactis MG1363. Microbiology 144:2885–2893 [CrossRef]
     [Google Scholar]
  33. Yamanaka K., Inouye M. 1997; Growth-phase-dependent expression of cspD, encoding a member of the CspA family in Escherichia coli. J Bacteriol 179:5126–5130
     [Google Scholar]
  34. Yamanaka K., Fang L., Inouye M. 1998; The CspA family in Escherichia coli: multiple gene duplication for stress adaptation. Mol Microbiol 27:247–255 [CrossRef]
     [Google Scholar]
/content/journal/micro/10.1099/00221287-145-11-3185
Loading
Analysis of the role of 7 kDa cold-shock proteins of Lactococcus lactis MG1363 in cryoprotection
Microbiology 145, 3185 (1999); https://doi.org/10.1099/00221287-145-11-3185
/content/journal/micro/10.1099/00221287-145-11-3185
/content/journal/micro/10.1099/00221287-145-11-3185
Loading

Data & Media loading...

Most cited Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error