Trehalose and Protein Stability

Nishant Kumar Jain1, Ipsita Roy1

1 Department of Biotechnology, National Institute of Pharmaceutical Education and Research (NIPER), S.A.S. Nagar, Punjab, India
Publication Name:  Current Protocols in Protein Science
Unit Number:  Unit 4.9
DOI:  10.1002/0471140864.ps0409s59
Online Posting Date:  February, 2010
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The role of osmolytes, and especially trehalose, in stabilizing proteins under stress conditions is now a widely accepted fact. The physical and chemical properties of trehalose, i.e., low chemical reactivity, nonreducing nature, high glass transition temperature, high affinity for water molecules, existence of a number of polymorphs, etc., make it uniquely suitable for stabilizing partially unfolded protein molecules and inhibiting protein aggregation. This article discusses the various adverse situations that protein molecules face, both within the cell and outside, leading to their aggregation and inactivation. The use of trehalose in stabilizing protein molecules and helping them retain their functionally active forms under such conditions is examined. The various theories and mechanisms used to explain the protective action of trehalose are briefly presented. The experimental tools that can be used to decipher the mechanism of aggregation and the role of trehalose are also discussed. Curr. Protoc. Protein Sci. 59:4.9.1‐4.9.12. © 2010 by John Wiley & Sons, Inc.

Keywords: anhydrobiosis; protein aggregation; protein stabilization

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Table of Contents

  • Introduction
  • Theories of Protein Stability with Respect to Trehalose
  • Desiccation/Lyophilization of Protein and Trehalose
  • Techniques Involved
  • Conclusion
  • Literature Cited
  • Tables
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Literature Cited

Literature Cited
   Attanasio, F., Cascio, C., Fisichella, S., Nicoletti, V.G., Pignataro, B., Savarino, A., and Rizzarelli, E. 2007. Trehalose effects on α‐crystallin aggregates. Biochem. Biophys. Res. Commun. 354:899‐905.
   Baptista, R.P., Cabral, J.M.S., and Melo, E.P. 2000. Trehalose delays the reversible but not the irreversible thermal denaturation of cutinase. Biotechnol. Bioeng. 70:699‐703.
   Béranger, F., Crozet, C., Goldsborough, A., and Lehmann, S. 2008. Trehalose impairs aggregation of PrPSc molecules and protects prion‐infected cells against oxidative damage. Biochem. Biophys. Res. Commun. 374:44‐48.
   Braun, A., Kwee, L., Labow, M.A., and Alsenz, J. 1997. Protein aggregates seem to play a key role among the parameters influencing the antigenicity of interferon alpha (IFN‐alpha) in normal and transgenic mice. Pharm. Res. 14:1472‐1478.
   Carninci, P., Nishiyama, Y., Westover, A., Itoh, M., Nagaoka, S., Sasaki, N., Okazaki, Y., Muramatsu, M., and Hayashizaki, Y. 1998. Thermostabilization and thermoactivation of thermolabile enzymes by trehalose and its application for the synthesis of full length cDNA. Proc. Natl. Acad. Sci. U.S.A. 95:520‐524.
   Carpenter, J.F., Crowe, L.M., and Crowe, J.H. 1987. Stabilization of phosphofructokinase with sugars during freeze‐drying: Characterization of enhanced protection in the presence of divalent cations. Biochim. Biophys. Acta 923:109‐115.
   Chen, L., Cao, L., Zhou, L., Jing, Y., Chen, Z., Deng, C., Shen, Y., and Chen, L. 2007. Trehalose as a good candidate for enriching full‐length cDNAs in cDNA library construction. J. Biotechnol. 127:402‐407.
   Davies, J.E., Sarkar, S., and Rubinsztein, D.C. 2006. Wild‐type PABPN1 is anti‐apoptotic and reduces toxicity of the oculopharyngeal muscular dystrophy mutation. Hum. Mol. Genet. 15:23‐31.
   Han, Y., Jin, B.S., Lee, S.B., Sohn, Y., Joung, J.W., and Lee, J.H. 2007. Effects of sugar additives on protein stability of recombinant human serum albumin during lyophilization and storage. Arch. Pharm. Res. 30:1124‐1131.
   Hédoux, A., Willart, J‐F., Paccou, L., Guinet, Y., Affouard, F., Lerbret, A., and Descamps, M. 2009. Thermostabilization mechanism of bovine serum albumin by trehalose. J. Phys. Chem. B 11:6119‐6126.
   Heikal, A., Box, K., Rothnie, A., Storm, J., Callaghan, R., and Allen, M. 2009. The stabilization of purified, reconstituted P‐glycoprotein by freeze drying with disaccharides. Cryobiology. 58:37‐44.
   Hulse, W.L., Forbes, R.T., Bonner, M.C., and Getrost, M. 2008. Do co‐spray dried excipients offer better lysozyme stabilisation than single excipients? Eur. J. Pharm. Sci. 33:294‐305.
   Jain, N.K. and Roy, I. 2008. Role of trehalose in moisture‐induced aggregation of bovine serum albumin. Eur. J. Pharm. Biopharm. 69:824‐834.
   Jain, N.K. and Roy, I. 2009. Effect of trehalose on protein structure. Protein Sci. 18:24‐36.
   Kawai, K. and Suzuki, T. 2007. Stabilizing effect of four types of disaccharide on the enzymatic activity of freeze‐dried lactate dehydrogenase: Step by step evaluation from freezing to storage. Pharm. Res. 24:1883‐1890.
   Liu, R., Barkhordarian, H., Emadi, S., Park, C.B., and Sierks, M.R. 2005. Trehalose differentially inhibits aggregation and neurotoxicity of beta amyloid 40 and 42. Neurobiol. Dis. 20:74‐81.
   López‐Díez, E.C. and Bone, S. 2004. The interaction of trypsin with trehalose: An investigation of protein preservation mechanisms. Biochim. Biophys. Acta 1673:139‐148.
   Lu, J., Wang, X.‐J., Liu, Y.‐X., and Ching, C.‐B. 2007. Thermal and FTIR investigation of freeze‐dried protein‐excipient mixtures. J. Therm. Anal. Calorim. 89:913‐919.
   Luo, Y., Li, W‐M., and Wang, W. 2008. Trehalose: Protector of antioxidant enzymes or reactive oxygen species scavenger under heat stress? Environ. Exper. Botan. 63:378‐384.
   Oriiand, Y. and Morita, M. 1977. Measurement of the pH of frozen buffer solutions by using pH indicators. J. Biochem. (Tokyo) 81:163‐168.
   Paz‐Alfaro, K.J., Ruiz‐Granadosb, Y.G., Uribe‐Carvajal, S., and Sampedroa, J.G. 2009. Trehalose‐mediated thermal stabilization of glucose oxidase from Aspergillus niger. J. Biotechnol. 141:130‐136.
   Qoronfleh, M.W., Hesterberg, L.K., and Seefeldt, M.B. 2007. Confronting high‐throughput protein refolding using high pressure and solution screens. Protein Expr. Purif. 55:209‐224.
   Roy, I. and Gupta, M.N. 2004. Freeze‐drying of proteins: Some emerging concerns. Biotechnol. Appl. Biochem. 39:165‐177.
   Saadati, Z. and Bordbar, A.K. 2008. Stability of β‐lactoglobulin a in the presence of sugar osmolytes estimated from their guanidinium chloride‐induced transition curves. Protein J. 27:455‐460.
   Santagapita, P.R., and Buera, M.P. 2008. Trehalose‐water‐salt interactions related to the stability of β‐galactosidase in supercooled media. Food Biophys. 3:87‐93.
   Singer, M.A. and Lindquist, S. 1998. Multiple effects of trehalose on protein folding in vitro and in vivo. Mol. Cell. 1:639‐648.
   Sola‐Penna, M. and Meyer‐Fernandes, J.R. 1996. Trehalose protects yeast pyrophosphatase against structural and functional damage induced by guanidinium chloride. Z. Naturforsch. C 51:160‐164.
   Sola‐Penna, M. and Meyer‐Fernandes, J.R. 1998. Stabilization against thermal inactivation promoted by sugars on enzyme structure and function: Why is trehalose more effective than other sugars? Arch. Biochem. Biophys. 360:10‐14.
   Sola‐Penna, M. and Meyer‐Fernandes, J.R. 2005. Trehalose and glycerol stabilize and renature yeast inorganic pyrophosphatase inactivated by very high temperatures. Arch. Biochem. Biophys. 444:52‐60.
   Spiess, A.N., Mueller, N., and Ivell, R. 2004. Trehalose is a potent PCR enhancer: Lowering of DNA melting temperature and thermal stabilization of taq polymerase by the disaccharide trehalose. Clin. Chem. 50:1256‐1259.
   Tanaka, M., Michida, Y., Niu, S., Ikeda, T., Jana, N.R., Doi, H., Kurosawa, M., Nekooki, M., and Nukina, N. 2004. Trehalose alleviates polyglutamine‐mediated pathology in a mouse model of Huntington's disease. Nat. Med. 10:148‐154.
   Zhang, Y., Ji, B., Ling, P., and Zhang, T. 2007. Trehalose and hyaluronic acid coordinately stabilized freeze‐dried pancreatic kininogenase. Eur. J. Pharm. Biopharm. 65:18‐25.
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