P-azidophenylglyoxal-epidermal growth factor: A photoactivatable affinity crosslinking reagent

Robert C. Miller, Stephen Planck, Bruce E. Magun

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

A photoaffinity derivative of highly purified 125I-labelled epidermal growth factor (125I-EGF) has been synthesized. The heterobifunctional crosslinking reagent p-azidophenylglyoxal (PAPG) was bound to arginine residues in 125I-EGF. PAPG-125I-EGF bound to EGF receptors on rat fibroblasts and human A431 epidermoid carcinoma cells in culture. An apparent decreased affinity of PAPG-125I-EGF for the EGF receptor is in accord with at least one arginine being at or near the EGF receptor binding site. The PAPG-125I-EGF:EGF receptor complexes on rat cells were internalized to the same extent as control EGF:receptor complexes. A431 cells treated with PAPG-125I-EGF were irradiated with ultraviolet light and the labelled proteins were analyzed by SDS-polyacrylamide gel electrophoresis. The 3 major labelled proteins had apparent molecular weights ranging from 75,000 to 200,000. Only the labelling of the 200,000-Mr protein was prevented by the addition of excess unlabelled EGF with the PAPG-125I-EGF. This molecular weight is in agreement with the reported size of the EGF receptor plus EGF. A protein with apparent molecular weight of 100,000 was labelled by 125I-EGF by an unknown mechanism which was dependent on the dose of UV light and blocked by the addition of excess unlabelled EGF.

Original languageEnglish (US)
Pages (from-to)529-539
Number of pages11
JournalJournal of Receptors and Signal Transduction
Volume3
Issue number4
DOIs
StatePublished - 1983
Externally publishedYes

Fingerprint

Cross-Linking Reagents
Epidermal Growth Factor
Epidermal Growth Factor Receptor
Molecular Weight
Molecular weight
Ultraviolet Rays
Arginine
Rats
Proteins
Fibroblasts
Electrophoresis
Cell culture
Ultraviolet radiation
Labeling
4-azidophenylglyoxal
Polyacrylamide Gel Electrophoresis
Squamous Cell Carcinoma
Cell Culture Techniques

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology
  • Pharmacology

Cite this

P-azidophenylglyoxal-epidermal growth factor : A photoactivatable affinity crosslinking reagent. / Miller, Robert C.; Planck, Stephen; Magun, Bruce E.

In: Journal of Receptors and Signal Transduction, Vol. 3, No. 4, 1983, p. 529-539.

Research output: Contribution to journalArticle

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