Epitope mapping of horseradish peroxidase with use of monoclonal antibodies

David Tipton, W. S. Walker, G. R. Schonbaum

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Nine mouse monoclonal antibodies (MAbs) to horseradish peroxidase (HRP) were used to develop an epitope map of the enzyme. The results of competitive binding assay indicated three distinct patterns of reactivity. Two groups of MAbs (I and III) recognized epitopes located in separate antigenic regions on the molecule; another (II) bound to sites that overlapped with epitopes in either region I or III. Further definition of these regions was obtained by analyzing the MAbs for their binding to isolated heme, other peroxidases and heme-containing proteins, and to denatured and apo-HRP. None of the group I MAbs bound heme, suggesting that this region was removed from the active site of the enzyme. All of the group II and III MAbs bound heme as well as the other peroxidases and heme-containing proteins, indicating that they recognized heme-associated epitopes at or near the active site. Only one MAb (2A2) in groups II and III bound to apo-HRP but not to denaturated HRP; it was also the only MAb in the entire panel that inhibited the catalytic activity of HRP. This suggests that the epitope recognized by 2A2 involves both the heme moiety and a conformationally dependent protein determinant near the active site.

Original languageEnglish (US)
Pages (from-to)319-330
Number of pages12
JournalHybridoma
Volume9
Issue number4
DOIs
StatePublished - Jan 1 1990

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Epitope Mapping
Horseradish Peroxidase
Heme
Monoclonal Antibodies
Epitopes
Peroxidases
Catalytic Domain
Proteins
Competitive Binding
Enzymes

All Science Journal Classification (ASJC) codes

  • Immunology
  • Genetics

Cite this

Epitope mapping of horseradish peroxidase with use of monoclonal antibodies. / Tipton, David; Walker, W. S.; Schonbaum, G. R.

In: Hybridoma, Vol. 9, No. 4, 01.01.1990, p. 319-330.

Research output: Contribution to journalArticle

Tipton, David ; Walker, W. S. ; Schonbaum, G. R. / Epitope mapping of horseradish peroxidase with use of monoclonal antibodies. In: Hybridoma. 1990 ; Vol. 9, No. 4. pp. 319-330.
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