Purification of Amyloid Protein AA Subspecies From Amyloid-Rich Human Tissues

  • Gunilla T. Westermark
  • Per Westermark
Part of the Methods in Molecular Biology™ book series (MIMB, volume 299)

Abstract

Protein AA, the major amyloid fibril protein in reactive (secondary) systemic amyloidosis is derived from the acute phase reactant liver-produced apolipoprotein serum AA (SAA) by proteolytic cleavage, usually in the C-terminal half of the 104 amino acid residues long precursor. The cleavage points in SAA vary between patients and the deposited protein AA is often quite heterogeneous. In this chapter, we describe methods to extract amyloid fibrils and to purify protein AA by sequential gel filtration. Further purification of subspecies of protein AA is best achieved by the use of differences in charge and chromatofocusing is described as the method of choice. Analytic methods include sodium dodecylsulfate polyacrylamide gel electrophoresis and analytic isoelectric focusing.

Key Words

econdary amyloidosis apolipoprotein fibril gel filtration isoelectric point protein fragment isoelectric focusing chromatofocusing polybuffer AA-subtypes amyloid extraction 

References

  1. 1.
    Benditt, E. P., Eriksen, N., Hermodson, M. A., and Ericsson, L. H. (1971) The major proteins of human and monkey amyloid substance: common properties including unusual N-terminal amino acid sequences. FEBS Lett. 19, 169–173.PubMedCrossRefGoogle Scholar
  2. 2.
    Westermark, G. T., Westermark, P., and Sletten, K. (1987) Amyloid fibril protein AA. Characterization of uncommon subspecies from a patient with rheumatoid arthritis. Lab. Invest. 57, 57–64.PubMedGoogle Scholar
  3. 3.
    Westermark, G. T., Sletten, K., and Westermark, P. (1989) Massive vascular AA-amyloidosis: A histologically and biochemically distinctive subtype of reactive systemic amyloidosis. Scand. J. Immunol. 30, 605–613.PubMedCrossRefGoogle Scholar
  4. 4.
    Westermark, G. T., Sletten, K., Grubb, A., and Westermark, P. (1990) AA-amy-loidosis. Tissue component-specific association of various protein AA subspecies and evidence of a fourth SAA gene product. Am. J. Path. 137, 377–383.PubMedGoogle Scholar
  5. 5.
    Uhlar, M. and Whitehead, A. S. (1999) Serum amyloid A, the major vertebrate acute-phase reactant. Eur. J. Biochem. 265, 501–523.PubMedCrossRefGoogle Scholar
  6. 6.
    Husby, G., Marhaug, G., Dowton, A., Sletten, K., and Sipe, J. D. (1994) Serum amyloid A (SAA): biochemistry, genetics and the pathogenesis of AA amyloidosis. Amyloid: Int. J. Exp. Clin. Invest. 1, 119–137.Google Scholar
  7. 7.
    Møyner, Sletten, Husby, G., and Natvig, J. (1980) An unusually large (83 amino acid residues) amyloid fibril protein AA from a patient with Walden-ström's macroglobulinaemia and amyloidosis. Scand. J. Immunol. 11, 549–554.PubMedCrossRefGoogle Scholar
  8. 8.
    Liepnieks, J. J., Kluve-Beckerman, and Benson, M. D. (1995) Characterization of amyloid A protein in human secondary amyloidosis: the predominant deposition of serum amyloid A1. Biochim. Biophys. Acta 1270, 81–86.PubMedGoogle Scholar
  9. 9.
    Levin, M., Franklin, E. C., Frangione, and Pras, M. (1972) The amino acid sequence of a major nonimmunoglobulin component of some amyloid fibrils. J. Clin. Invest. 51, 2773–2776.PubMedCrossRefGoogle Scholar
  10. 10.
    Sletten, K. and Husby, G. (1974) The complete amino-acid sequence of non-immunoglobulin amyloid fibril protein AS in rheumatoid arthritis. Eur. J. Biochem. 41, 117–125.PubMedCrossRefGoogle Scholar
  11. 11.
    Shirahama, T. and Cohen, A. S. (1967) High-resolution electron microscopic analysis of the amyloid fibril. J. Cell. Biol. 33, 679–708.PubMedCrossRefGoogle Scholar
  12. 12.
    Pras, M., Schubert, M., Zucker-Franklin, D., Rimon, A., and Franklin, E. (1968) The characterization of soluble amyloid prepared in water. J. Clin. Invest. 47, 924–933.PubMedCrossRefGoogle Scholar
  13. 13.
    Skinner, M., Shirahama, T., Cohen, A. S., and Deal, L. (1983) The association of amyloid P-component (AP) with the amyloid fibril: an updated method for amyloid fibril protein isolation. Prep. Biochem. 12, 461–476.CrossRefGoogle Scholar
  14. 14.
    Westermark, P. (1982) The heterogeneity of protein AA in secondary (reactive) systemic amyloidosis. Biochim. Biophys. Acta 701, 19–23.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press Inc. 2005

Authors and Affiliations

  • Gunilla T. Westermark
    • 1
  • Per Westermark
    • 2
  1. 1.Department of Biomedicine and Surgery, Division of Cell BiologyLinköping UniversityLinköpingSweden
  2. 2.Department of Genetics and Pathology, Rudbeck LaboratoryUppsala UniversityUppsalaSweden

Personalised recommendations