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Methods for the Preparation and Crystallization of Fab Fragments in Complex with Peptide Antigens Derived from N. meningitidis Outer Membrane Proteins

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Part of the Methods in Molecular Medicine™ book series (MIMM,volume 66)

Abstract

An understanding of the molecular basis for the recognition of outer-membrane proteins (OMPs) by antibody is an important goal in the development of a more rational approach to vaccine design. X-ray crystallography has been outstandingly successful in delineating the detailed chemical interactions that are responsible for the high affinity and high selectivity of antibody-antigen interactions (1). Although a number of X-ray structures of OMPs have now been reported (e.g., 2), determination of the structure of a novel OMP is far from routine. Furthermore, it is more useful to know the structural basis for molecular recognition of a particular antigen by antibody than the structure of the cognate antigen alone. For this reason, work in this area has concentrated on studying the structures of antigen in complex with antibody Fab fragments. In practice, this requires the synthesis of a short peptide or oligosaccharide that binds to the antibody in question and then determination of the structure of the bound antigen by X-ray crystallography. Clearly, this has the limitation that only well-defined continuous epitopes can be studied in this way, but fortunately this is frequently the case with monoclonal antibodies (MAbs) that have been prepared against meningococcal OMPs. This type of approach has also been used to study the binding of the polysaccharide O-antigen from Shigella flexneri (3), and in principle it could be applied to meningococcal polysaccharide antigens if suitable small oligosaccharides could be synthesised in milligram quantities.

Keywords

  • Sodium Azide
  • Shigella Flexneri
  • Precipitant Concentration
  • Column Buffer
  • Ultrafiltration Cell

These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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© 2001 Humana Press Inc., Totowa, NJ

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Collins, R.F., Derrick, J.P. (2001). Methods for the Preparation and Crystallization of Fab Fragments in Complex with Peptide Antigens Derived from N. meningitidis Outer Membrane Proteins. In: Pollard, A.J., Maiden, M.C. (eds) Meningococcal Vaccines. Methods in Molecular Medicine™, vol 66. Humana Press. https://doi.org/10.1385/1-59259-148-5:121

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  • DOI: https://doi.org/10.1385/1-59259-148-5:121

  • Publisher Name: Humana Press

  • Print ISBN: 978-0-89603-801-1

  • Online ISBN: 978-1-59259-148-0

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