Abstract
Protein aggregation into amyloid conformations is associated with more than 50 different human disorders. Recent studies demonstrate that the expression in bacteria of amyloid proteins results in the formation of intracellular aggregates structurally related to those underlying human diseases. The ease with which prokaryotic organisms can be genetically and biochemically manipulated makes them useful systems for studying how and why protein aggregates inside the cell, providing a tractable environment to rationally model in vivo amyloid formation. In this chapter we present an overview of the methods used to characterize the kinetic, structural, and functional properties of amyloid-like bacterial intracellular aggregates and how they can be employed to screen for lead compounds that might modulate amyloid deposition.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Invernizzi G, Papaleo E, Sabate R et al (2012) Protein aggregation: mechanisms and functional consequences. Int J Biochem Cell Biol 44:1541–1554
Calamai M, Kumita JR, Mifsud J et al (2006) Nature and significance of the interactions between amyloid fibrils and biological polyelectrolytes. Biochemistry 45:12806–12815
Fernandez-Busquets X, de Groot NS, Fernandez D et al (2008) Recent structural and computational insights into conformational diseases. Curr Med Chem 15:1336–1349
Nelson R, Eisenberg D (2006) Recent atomic models of amyloid fibril structure. Curr Opin Struct Biol 16:260–265
Ventura S, Villaverde A (2006) Protein quality in bacterial inclusion bodies. Trends Biotechnol 24:179–185
de Groot NS, Sabate R, Ventura S (2009) Amyloids in bacterial inclusion bodies. Trends Biochem Sci 34:408–416
Sabate R, de Groot NS, Ventura S (2010) Protein folding and aggregation in bacteria. Cell Mol Life Sci 67:2695–2715
Garcia-Fruitos E, Sabate R, de Groot NS et al (2011) Biological role of bacterial inclusion bodies: a model for amyloid aggregation. FEBS J 278:2419–2427
Carrio M, Gonzalez-Montalban N, Vera A et al (2005) Amyloid-like properties of bacterial inclusion bodies. J Mol Biol 347:1025–1037
Morell M, Bravo R, Espargaro A et al (2008) Inclusion bodies: specificity in their aggregation process and amyloid-like structure. Biochim Biophys Acta 1783:1815–1825
Dasari M, Espargaro A, Sabate R et al (2011) Bacterial inclusion bodies of Alzheimer’s disease beta-amyloid peptides can be employed to study native-like aggregation intermediate states. Chembiochem 12:407–423
de Groot NS, Ventura S (2006) Protein activity in bacterial inclusion bodies correlates with predicted aggregation rates. J Biotechnol 125:110–113
de Groot NS, Aviles FX, Vendrell J et al (2006) Mutagenesis of the central hydrophobic cluster in Abeta42 Alzheimer’s peptide. Side-chain properties correlate with aggregation propensities. FEBS J 273:658–668
Villar-Pique A, Espargaro A, Sabate R et al (2012) Using bacterial inclusion bodies to screen for amyloid aggregation inhibitors. Microb Cell Fact 11:55
Villar-Pique A, de Groot NS, Sabate R et al (2012) The effect of amyloidogenic peptides on bacterial aging correlates with their intrinsic aggregation propensity. J Mol Biol 421:270–281
Jahn TR, Radford SE (2008) Folding versus aggregation: polypeptide conformations on competing pathways. Arch Biochem Biophys 469:100–117
Garcia-Fruitos E, Gonzalez-Montalban N, Morell M et al (2005) Aggregation as bacterial inclusion bodies does not imply inactivation of enzymes and fluorescent proteins. Microb Cell Fact 4:27
Shimomura O, Johnson FH, Saiga Y (1962) Extraction, purification and properties of aequorin, a bioluminescent protein from the luminous hydromedusan, Aequorea. J Cell Comp Physiol 59:223–239
Tsien RY (1998) The green fluorescent protein. Annu Rev Biochem 67:509–544
Waldo GS, Standish BM, Berendzen J et al (1999) Rapid protein-folding assay using green fluorescent protein. Nat Biotechnol 17:691–695
Belli M, Ramazzotti M, Chiti F (2011) Prediction of amyloid aggregation in vivo. EMBO Rep 12:657–663
Castillo V, Grana-Montes R, Sabate R et al (2011) Prediction of the aggregation propensity of proteins from the primary sequence: aggregation properties of proteomes. Biotechnol J 6:674–685
Guidolin D, Agnati LF, Albertin G et al (2012) Bioinformatics aggregation predictors in the study of protein conformational diseases of the human nervous system. Electrophoresis 33:3669–3679
Conchillo-Sole O, de Groot NS, Aviles FX et al (2007) AGGRESCAN: a server for the prediction and evaluation of “hot spots” of aggregation in polypeptides. BMC Bioinformatics 8:65
Woulfe J (2008) Nuclear bodies in neurodegenerative disease. Biochim Biophys Acta 1783:2195–2206
Kopito RR (2000) Aggresomes, inclusion bodies and protein aggregation. Trends Cell Biol 10:524–530
Lindner AB, Madden R, Demarez A et al (2008) Asymmetric segregation of protein aggregates is associated with cellular aging and rejuvenation. Proc Natl Acad Sci U S A 105:3076–3081
Kim W, Kim Y, Min J et al (2006) A high-throughput screen for compounds that inhibit aggregation of the Alzheimer’s peptide. ACS Chem Biol 1:461–469
Martinez-Alonso M, Vera A, Villaverde A (2007) Role of the chaperone DnaK in protein solubility and conformational quality in inclusion body-forming Escherichia coli cells. FEMS Microbiol Lett 273:187–195
Vera A, Gonzalez-Montalban N, Aris A et al (2007) The conformational quality of insoluble recombinant proteins is enhanced at low growth temperatures. Biotechnol Bioeng 96:1101–1106
de Groot NS, Ventura S (2006) Effect of temperature on protein quality in bacterial inclusion bodies. FEBS Lett 580:6471–6476
Espargaro A, Sabate R, Ventura S (2012) Thioflavin-S staining coupled to flow cytometry. A screening tool to detect in vivo protein aggregation. Mol Biosyst 8:2839–2844
Rajan RS, Illing ME, Bence NF et al (2001) Specificity in intracellular protein aggregation and inclusion body formation. Proc Natl Acad Sci U S A 98:13060–13065
Hart RA, Rinas U, Bailey JE (1990) Protein composition of Vitreoscilla hemoglobin inclusion bodies produced in Escherichia coli. J Biol Chem 265:12728–12733
Wang L, Maji SK, Sawaya MR et al (2008) Bacterial inclusion bodies contain amyloid-like structure. PLoS Biol 6:e195
Cano-Garrido O, Rodriguez-Carmona E, Diez-Gil C et al (2013) Supramolecular organization of protein-releasing functional amyloids solved in bacterial inclusion bodies. Acta Biomater 9:6134–6142
Hubbard SJ (1998) The structural aspects of limited proteolysis of native proteins. Biochim Biophys Acta 1382:191–206
Kong J, Yu S (2007) Fourier transform infrared spectroscopic analysis of protein secondary structures. Acta Biochim Biophys Sin (Shanghai) 39:549–559
Tycko R (2006) Solid-state NMR as a probe of amyloid structure. Protein Pept Lett 13:229–234
Denizot F, Lang R (1986) Rapid colorimetric assay for cell growth and survival. Modifications to the tetrazolium dye procedure giving improved sensitivity and reliability. J Immunol Methods 89:271–277
Wasmer C, Benkemoun L, Sabate R et al (2009) Solid-state NMR spectroscopy reveals that E. coli inclusion bodies of HET-s(218–289) are amyloids. Angew Chem Int Ed Engl 48:4858–4860
Garrity SJ, Sivanathan V, Dong J et al (2010) Conversion of a yeast prion protein to an infectious form in bacteria. Proc Natl Acad Sci U S A 107:10596–10601
Espargaro A, Villar-Pique A, Sabate R et al (2012) Yeast prions form infectious amyloid inclusion bodies in bacteria. Microb Cell Fact 11:89
Liebman SW, Derkatch IL (1999) The yeast [PSI+] prion: making sense of nonsense. J Biol Chem 274:1181–1184
Tanaka M, Weissman JS (2006) An efficient protein transformation protocol for introducing prions into yeast. Methods Enzymol 412:185–200
Tanaka M (2010) A protein transformation protocol for introducing yeast prion particles into yeast. Methods Enzymol 470:681–693
Chernoff YO, Lindquist SL, Ono B et al (1995) Role of the chaperone protein Hsp104 in propagation of the yeast prion-like factor [psi+]. Science 268:880–884
Acknowledgments
This work was supported by grants BFU2010-14901 from Ministerio de Ciencia e Innovación (Spain), 2009-SGR-760 from AGAUR (Generalitat de Catalunya). S.V. has been granted an ICREA Academia award (ICREA).
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Science+Business Media New York
About this protocol
Cite this protocol
Villar-Pique, A., Navarro, S., Ventura, S. (2015). Characterization of Amyloid-Like Properties in Bacterial Intracellular Aggregates. In: García-Fruitós, E. (eds) Insoluble Proteins. Methods in Molecular Biology, vol 1258. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2205-5_6
Download citation
DOI: https://doi.org/10.1007/978-1-4939-2205-5_6
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2204-8
Online ISBN: 978-1-4939-2205-5
eBook Packages: Springer Protocols