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Biochemical implications from the variable gene-sequences of an anti-cytochrome-c antibody and crystallographic characterization of its antigen-binding fragment in free and antigen-complexed forms

Academic Article
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Overview

authors

  • Mylvaganam, S. E.
  • Paterson, Y.
  • Kaiser, K.
  • Bowdish, K.
  • Getzoff, Elizabeth

publication date

  • September 1991

journal

  • Journal of Molecular Biology  Journal

abstract

  • To study the nature of antibody-antigen interactions, we have determined the variable gene sequences of the anti-cytochrome c immunoglobulin G1 (IgG1) monoclonal antibody E8, and obtained diffraction-quality crystals of the E8 antigen-binding fragment (Fab), both free and bound to its antigen, horse cytochrome c. The FabE8 crystals belong to space group P21 with unit cell dimensions of a = 45.0 A, b = 85.1 A, c = 63.3 A and beta = 105.5 degrees, have one FabE8 molecule per asymmetric unit and diffract to at least 2.1 A resolution. Crystals of the FabE8-cytochrome c complex belong to space group P212121 with unit cell dimensions of a = 84.3 A, b = 73.3 A and c = 94.9 A, accommodate one complex per asymmetric unit and diffract to 2.4 A resolution. In the nucleotide-derived amino acid sequences, the light-chain variable domain (VL) but not the heavy-chain variable domain (VH) of E8 is nearly identical to that of the anti-lysozyme antibody D1.3, differing by only five amino acid residues. Only one of these interacts with lysozyme in the D1.3-lysozyme crystal structure. Six negative and four positive charges in the VH complementarity determining regions of E8 complement four positive and three negative charges in the E8 epitope on cytochrome c. These data suggest that only a subset of the residues in an antibody-protein interface may be critical for binding and that the VH may play a dominant role in antigenic recognition.

subject areas

  • Amino Acid Sequence
  • Animals
  • Antibodies, Monoclonal
  • Antigen-Antibody Complex
  • Antigen-Antibody Reactions
  • Base Sequence
  • Cell Line
  • Crystallization
  • Cytochrome c Group
  • Genes, Immunoglobulin
  • Immunoglobulin Fab Fragments
  • Immunoglobulin G
  • Immunoglobulin Variable Region
  • Mice
  • Mice, Inbred BALB C
  • Molecular Sequence Data
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Research

keywords

  • ANTIBODY SEQUENCE
  • CRYSTALLIZATION
  • CYTOCHROME-C-ANTIBODY COMPLEX
  • IMMUNOGLOBULIN
  • X-RAY DIFFRACTION
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Identity

International Standard Serial Number (ISSN)

  • 0022-2836

Digital Object Identifier (DOI)

  • 10.1016/0022-2836(91)80066-4

PubMed ID

  • 1656053
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Additional Document Info

start page

  • 455

end page

  • 462

volume

  • 221

issue

  • 2

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