Document Type : Original Article
Authors
1 Department of Biochemistry, Payam Noor University, Tehran, Iran
2 Department of Clinical Biochemistry, Tarbiat Modares University, Tehran, Iran
3 Young Researchers and Elite Club, Tehran Medical Sciences Branch, Islamic Azad University, Tehran, Iran
Abstract
Background: Molecular chaperon-like activity for protein refolding was studied using nanogel chitosan-myristic acid (CMA) and the protein neuroserpin (NS), a member of the serine proteinase inhibitor superfamily (serpin).
Materials and Methods: Recombinant his-tag fusion NS was expressed in Escherichia coli. For confirmation of refolding of the purified NS, structural analysis was performed by circular dichroism and spectrofluorometric along with its inhibitory activity, which was assayed by single-chain tissue plasminogen activator. For evaluating NS aggregation during preparation, the samples were separated on a 7.5% (w/v) nondenaturing polyacrylamide gel electrophoresis. MA and chitosan covalently join together by the formation of amide linkages through the 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide-mediated reaction. The morphology and size of the prepared CM nanogel were characterized by transmission electron microscopy and scanning electron microscopy.
Results: Heating at different temperatures (25°C, 37°C, 45°C, 65°C, 80°C) results in a further rise in β-structures accompanied by a fall of helices and no significant change in random coils. Structural changes in NS in the presence of CMA nanogel were less than that in the absence of CMA nanogel. Mater nanogel effectively prevented aggregation of NS during temperature induced protein refolding by the addition of cyclodextrins. The nanogel activity resembled the host-guest chaperon activity.
Conclusion: These conditions, called conformational disorders, include Alzheimer's, Parkinson's, Huntington's disease, the transmissible spongiform encephalopathies, prion diseases, and dementia. Nanogels can be useful in recovery of the structural normality of proteins in these diseases.
Keywords
1. |
Osterwalder T, Contartese J, Stoeckli ET, Kuhn TB, Sonderegger P. Neuroserpin, an axonally secreted serine protease inhibitor. EMBO J 1996;15:2944-53. [PUBMED] |
2. |
Schrimpf SP, Bleiker AJ, Brecevic L, Kozlov SV, Berger P, Osterwalder T, et al. Human neuroserpin (PI12): CDNA cloning and chromosomal localization to 3q26. Genomics 1997;40:55-62. [PUBMED] |
3. |
Krueger SR, Ghisu GP, Cinelli P, Gschwend TP, Osterwalder T, Wolfer DP, et al. Expression of neuroserpin, an inhibitor of tissue plasminogen activator, in the developing and adult nervous system of the mouse. J Neurosci 1997;17:8984-96. [PUBMED] |
4. |
Whisstock J, Skinner R, Lesk AM. An atlas of serpin conformations. Trends Biochem Sci 1998;23:63-7. [PUBMED] |
5. | |
6. |
Dean RT, Fu S, Stocker R, Davies MJ. Biochemistry and pathology of radical-mediated protein oxidation. Biochem J 1997;324:1-18. [PUBMED] |
7. |
Forster MJ, Dubey A, Dawson KM, Stutts WA, Lal H, Sohal RS. Age-related losses of cognitive function and motor skills in mice are associated with oxidative protein damage in the brain. Proc Natl Acad Sci U S A 1996;93:4765-9. [PUBMED] |
8. |
Aksenov MY, Aksenova MV, Butterfield DA, Geddes JW, Markesbery WR. Protein oxidation in the brain in Alzheimer′s disease. Neuroscience 2001;103:373-83. [PUBMED] |
9. |
Varadarajan S, Yatin S, Aksenova M, Butterfield DA. Review: Alzheimer′s amyloid beta-peptide-associated free radical oxidative stress and neurotoxicity. J Struct Biol 2000;130:184-208. [PUBMED] |
10. |
Mohsenifar A, Lotfi AS, Ranjbar B, Allameh A, Zaker F, Hasani L, et al. A study of the oxidation-induced conformational and functional changes in neuroserpin. Iran Biomed J 2007;11:41-6. [PUBMED] |
11. |
Telling GC, Parchi P, DeArmond SJ, Cortelli P, Montagna P, Gabizon R, et al. Evidence for the conformation of the pathologic isoform of the prion protein enciphering and propagating prion diversity. Science 1996;274:2079-82. [PUBMED] |
12. |
Pan KM, Baldwin M, Nguyen J, Gasset M, Serban A, Groth D, et al. Conversion of alpha-helices into beta-sheets features in the formation of the scrapie prion proteins. Proc Natl Acad Sci U S A 1993;90:10962-6. [PUBMED] |
13. |
Prusiner SB, McKinley MP, Bowman KA, Bolton DC, Bendheim PE, Groth DF, et al. Scrapie prions aggregate to form amyloid-like birefringent rods. Cell 1983;35:349-58. [PUBMED] |
14. |
Agnihotri SA, Mallikarjuna NN, Aminabhavi TM. Recent advances on chitosan-based micro-and nanoparticles in drug delivery. J Control Release 2004;100:5-28. [PUBMED] |
15. |
Suh JK, Matthew HW. Application of chitosan-based polysaccharide biomaterials in cartilage tissue engineering: A review. Biomaterials 2000;21:2589-98. [PUBMED] |
16. | |
17. | |
18. | |
19. | |
20. |
Kelly SM, Jess TJ, Price NC. How to study proteins by circular dichroism. Biochim Biophys Acta 2005;1751:119-39. [PUBMED] |
21. | |
22. |
Schatz C, Viton C, Delair T, Pichot C, Domard A. Typical physicochemical behaviors of chitosan in aqueous solution. Biomacromolecules 2003;4:641-8. [PUBMED] |
23. | |
24. |
Popa-Nita S, Alcouffe P, Rochas C, David L, Domard A. Continuum of structural organization from chitosan solutions to derived physical forms. Biomacromolecules 2010;11:6-12. [PUBMED] |
25. | |
26. | |
27. |
Chen XG, Lee CM, Park HJ. O/W emulsification for the self-aggregation and nanoparticle formation of linoleic acid-modified chitosan in the aqueous system. J Agric Food Chem 2003;51:3135-9. [PUBMED] |
28. | |
29. |
Yoshioka H, Nonaka K, Fukuda K, Kazama S. Chitosan-derived polymer-surfactants and their micellar properties. Biosci Biotechnol Biochem 1995;59:1901-4. [PUBMED] |