Abstract
Members of the cyclophilin (Cyp) family are known to function as co-chaperones, interacting with chaperones such as heat shock protein 90, and perform important roles in protein folding under high temperature stress. In addition, they have been isolated from a wide range of organisms. However, there have been no reports on the functions of algal Cyps under other stress conditions. To study the functions of the cDNAGjCyp-1 isolated from the red alga (Griffithsia japonica), a recombinant GjCyp-1 containing a hexahistidine tag at the amino-terminus was constructed and expressed inEscherichia coli. Most of the gene product expressed inE. coli was organized as aggregate insoluble particles known as inclusion bodies. Thus, the optimal time, temperature, and concentration ofl(+)-arabinose for expressing the soluble and nonaggregated form of GjCyp-1 inE. coli were examined. The results indicate that the induction of Cyp, at 0.2%l(+)-arabinose for 2 h at 25°C, had a marked effect on the yield of the soluble and active form of the co-chaperone as PPlase. An expressed fusion protein, H6GjCyp-1, maintained the stability ofE. coli proteins up to-75°C. In a functional bioassay of the recombinant H6GjCyp-1, the viability ofE. coli cells overexpressing H6GjCyp-1 was compared to that of cells not expressing H6GjCyp-1 at −75°C. For all the cycles of a freeze/thaw treatment, a significant increase in viability was observed in theE. coli cells overexpressing H6GjCyp-1. The results of the GjCyp-1 bioassays, as well asin vitro studies, strongly suggest that the algal Cyp confers freeze tolerance toE. coli.
Similar content being viewed by others
References
Antikainen, M. and M. Griffith (1997) Antifreeze protein accumulation in freezing-tolerant cereals.Physiol. Plant. 99: 423–432.
Boothe, J. G., F. D. Sonnichsen, M. D. de Beus, and A. M. Johnson-Flanagan (1997) Purification, characterization, and structural analysis of a plant low-temperature-induced protein.Plant Physiol. 113: 367–376.
Ndong, C., F. Ouellet, M. Houde, and F. Sarhan (1997) Gene expression during cold acclimation in strawberry.Plant Cell Physiol. 38: 863–870.
Tiku, P. E., A. Y. Gracey, A. I. Macartney, R. J. Beynon, and A. R. Cossins (1996) Cold-induced expression of delta 9-desaturase in carp by transcriptional and posttranscriptional mechanisms.Science 271: 815–818.
Yamashita, M., N. Ojima, and T. Sakamoto (1996) Molecular cloning and cold-inducible gene expression of ferritin H subunit isoforms in rainbow trout cells.J. Biol. Chem. 271: 26908–26913.
Pearce, R. S. (1999) Molecular analysis of acclimation to cold.Plant Growth Regul. 29: 47–76.
Arora, R., L. J. Rowland, and G. R. Panta (1997) Chill-responsive dehydrins in blueberry: Are they associated with cold hardiness or dormancy transitions?Physiol. Plant. 101: 8–16.
Ferullo, J. M., L. P. Vezina, J. Rail, S. Laberge, P. Nadeau, and Y. Castonguay (1997) Differential accumulation of two glycine-rich proteins during cold-acclimation alfalfa.Plant Mol. Biol 33: 625–633.
Sokolova I. M. and H. O. Pörtner (2003) Metabolic plasticity and critical temperatures for aerobic scope in a eurythermal marine invertebrate (Littorina saxatilis, Gastropoda: Littorinidae) from different latitudes.J. Exp. Biol. 206: 195–207.
Zhu, B., T. H. H. Chen, and P. H. Li (1993) Expression of an ABA-responsive osmotin-like gene during the induction of freezing tolerance inSolanum commersonii.Plant Mol. Biol. 21: 729–735.
Lewis, J. G., R. P. Learmonth, and K. Watson (1995) Induction of heat, freezing and salt tolerance by heat and salt shock inSaccharomyces cerevisiae.Microbiology 141: 687–694.
Queitsch, C., S. W. Hong, E. Vierling, and S. Lindquist (2000) Heat shock protein 101 plays a crucial role in thermotolerance in Arabidopsis.Plant Cell 12: 479–492.
Boston, R. S., P. V. Viitanen, and E. Vierling (1996) Molecular chaperones and protein folding in plants.Plant Mol. Biol. 32: 191–222.
Cho, E. K., Y. K. Lee, and C. B. Hong (2005) A cyclophilin fromGriffithsia japonica has thermoprotective activity and is affected by CsA.Mol. Cells 20: 142–150.
Luan, S., W. S. Lane, and S. L. Schreiber (1994) pCyP B: a chloroplast-localized, heat shock-responsive cyclophilin from fava bean.Plant Cell 6: 885–892.
Marivet, J., M. Margis-Pinheiro, P. Frendo, and G. Burkard (1994) Bean cyclophilin gene expression during plant development and stress conditions.Plant Mol. Biol. 26: 1181–1189.
Marivet, J., M. Margispinheiro, P. Frendo, and G. Burkard (1995) DNA sequence analysis of a cyclophilin gene from maize: developmental expression and regulation by salicylic acid.Mol. Gen. Genet. 247: 222–228.
Mark, P. J., B. K. Ward, P. Kumar, H. Lahooti, R. F. Minchin, and T. Ratajczak (2001) Human cyclophilin 40 is a heat shock protein that exhibits altered intracellular localization following heat shock.Cell Stress Chaperones 6: 59–70.
Clubb, R. T., S. B. Ferguson, C. T. Walsh, and G. Wagner (1994) Three-dimensional solution structure ofEscherichia coli periplasmic cyclophilin.Biochemistry 33: 2751–2772.
Fejzo, J., F. A. Etzkorn, R. T. Clubb, Y. Shi, C. T. Walsh, and G. Wagner (1994) The mutantEscherichia coli F112W cyclophilin binds cyclosporin A in nearly identical conformation as human cyclophilin.Biochemistry 33: 5711–5720.
Lee, Y. K., C. B. Hong, Y. Suh, and I. K. Lee (2002) A cDNA clone for cyclophilin fromGriffithsia japonica and phylogenetic analysis of cyclophilins.Mol. Cells 13: 12–20.
English, T. E. and K. B. Storey (2003) Freezing and anoxia stresses induce expression of metallothionein in the foot muscle and hepatopancreas of the marine gastropodLittorina littorea.J. Exp. Biol. 206: 2517–2524.
Larade, K., A. Nimigan, and K. B. Storey (2001) Transcription pattern of ribosomal protein L26 during anoxia exposure inLittorina littorea.J. Exp. Zool. 290: 759–768.
Menally, J. D., S. B. Wu, C. M. Sturgeon, and K. B. Storey (2002) Identification and characterization of a novel freezing inducible gene,li16, in the wood frogRana sylvatica.FASEB J. 16: 902–904.
Kim, K. P., M. K. Joe, and C. B. Hong (2004) Tobacco small heat-shock protein, NtHSP18.2, has broad substrate range as a molecular chaperone.Plant Sci. 167: 1017–1025.
Bradford, M. M. (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding.Anal. Biochem. 72: 248–254.
Sambrook, J., E. F. Fritsch, and T. Maniatis (1989)Molecular Cloning: A Laboratory Manual. 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA.
Duina, A. A., J. A. Marsh, R. B. Kurtz, H. J. Chang S. Lindquist, and R. F. Gaber (1998) The peptidyl-prolyl isomerase domain of the CyP-40 cyclophilin homolog Cpr7 is not required to support growth or glucocorticoid receptor activity inSaccharomyces cerevisiae.J. Biol. Chem. 273: 10819–10822.
Liu, X. D., K. A. Morano, and D. J. Thiele (1999) The yeast Hsp110 family member, Ssel, is an Hsp90 cochaperone.J. Biol. Chem. 274: 26654–26660.
Weisman, R., J. Creanor, and P. Fantes (1996) A multicopy suppressor of a cell cycle defect inSchizosaccharomyces pombe encodes a heat shock-inducible 40 kDa cyclophilin-like protein.EMBO J. 15: 447–456.
Küllertz, G., A. Liebau, P. Rücknagel, A Schierhorn, B. Diettrich, G. Fischer, and M. Luckner (1999) Stress-induced expression of cyclophilins in proembryonic masses ofDigitalis lanata does not protect against freezing/thawing stress.Planta 208: 599–605.
Guy C. L., D. Haskell, and Q. B. Li (1998) Association of proteins with the stress 70 molecular chaperones at low temperature: evidence for the existence of cold labile proteins in spinach.Cryobiology 36: 301–314.
Sung, D. Y., E. Vierling, and C. L. Guy (2001) Comprehensive expression profile analysis of the Arabidopsis Hsp70 gene family.Plant Physiol. 126: 789–800.
Lopez-Matas, M. A., P. Nuñez, A Soto, I. Allona, R. Casado, C. Collada, M. A. Guevara, C. Aragoncillo, and L. Gomez (2004) Protein cryoprotective activity of a cytosolic small heat shock protein that accumulates constitutively in chestnut stems and is up-regulated by low and high temperatures.Plant Physiol 134: 1708–1717.
Place, S. P. and G. E. Hofmann (2005) Temperature differentially affects adenosine triphosphatase activity in Hsc70 orthologs from Antarctic and New Zealand notothenioid fishes.Cell Stress Chaperones 10: 104–113.
Walker, D. C., H. S. Girgis, and T. R. Klaenhammer (1999) ThegroESL chaperone operon ofLactobacillus johnsonii.Appl. Environ. Microbiol. 65: 3033–3041.
Chow, K. C. and W. L. Tung (1998) Overexpression ofdnaK/dnaJ andgroEL confers freeze tolerance toEscherichia coli.Biochem. Biophys. Res. Commun. 253: 502–505.
Tesic, M., J. A. Marsh, S. B. Cullinan, and R. F. Gaber (2003) Functional interactions between Hsp90 and the co-chaperones Cns1 and Cpr7 inSaccharomyces cerevisiae.J. Biol. Chem. 278: 32692–32701.
Edvardsson, A., S. Eshaghi, A. V. Vener, and B. Andersson (2003) The major peptidyl-prolyl isomerase activity in thylakoid lumen of plant chloroplasts belongs to a novel cyclophilin TLP20.FEBS Lett. 542: 137–141.
Kern, G., D. Kern, F. X. Schmid, and G Fischer (1994) Reassessment of the putative chaperone function of prolylcis/ trans-isomerase.FEBS Lett 348: 145–148.
Rutherford, S. L. and C. S. Zuker (1994) Protein folding and the regulation of signaling pathways.Cell 79: 1129–1132.
Kinoshita, T. and K. I. Shimazaki (1999) Characterization of cytosolic cyclophilin from guard cells ofVicia jaba L.Plant Cell Physiol. 40: 53–59.
Schneider, H., N. Charara, R. Schmitz, S. Wehrli, V. Mikol, M. G. Zurini, V. F. Quesniaux, and N. R. Movva (1994) Human cyclophilin C: primary structure tissue distribution, and determination of binding specificity for cyclosporins.Biochemistry 33: 8218–8224.
Kim, J. E., E. J. Kim, W. J. Rhee, and T. H. Park (2005) Enhanced production of recombinant protein inEscherichia coli using silkworm hemolymph.Biotechnol. Bioprocess Eng. 10: 353–356.
Shin, E. J., S. L. Park, S. J. Jeon, J. W. Lee, Y. T. Kim, Y. H. Kim, and S. W. Nam (2006) Effect of molecular chaperones on the soluble expression of alginate lyase inE. coli.Biotechnol Bioprocess Eng. 11: 414–419.
Kim, H. and I. H. Kim (2005) Refolding of fusion ferritin by gel filtration chromatography (GFC).Biotechnol. Bioprocess Eng. 10: 500–504.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Cho, E.K. Enhanced tolerance against freezing stress inEscherichia coli cells expressing an algal cyclophilin gene. Biotechnol. Bioprocess Eng. 12, 502–507 (2007). https://doi.org/10.1007/BF02931347
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF02931347