Pseudomonas exotoxin A-epidermal growth factor (EGF) mutant chimeric protein as an indicator for identifying amino acid residues important in EGF-receptor interaction

Her Shyong Shiah, Tzong Yueh Chen, Chi Ming Chang, Judy T. Chow, Hsing-Jien Kung, Jaulang Hwang

Research output: Contribution to journalArticle

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Abstract

Epidermal growth factor (EGF) was fused to the carboxyl end of a modified pseudomonas exotoxin A that has its toxin binding domain deleted. This chimeric toxin designated as PE(ΔIa)-EGF kills A431 cells through the EGF receptor-mediated pathway. In this study, we used a random mutagenesis approach to make point mutations on EGF, followed by replacing the wild type EGF in PE(ΔIa)-EGF with these EGF mutants. We have constructed 14 different PE(ΔIa)-EGFmutants, and examined their EGF receptor binding activity as well as their cytotoxicity to A431 cells. Our results showed that individual mutations of Val19 to Glu and Val34 to Asp in the EGF domain of PE(ΔIa)-EGFmutants resulted in an increase in the binding affinity to EGF receptor and cytotoxicity to A431 cells. On the other hand, individual mutations of His16 to Asp and Gly18 to Ala in the EGF domain of PE(ΔIa)-EGFmutants lead to a decrease in the binding affinity to EGF receptor and cytotoxicity to A431 cells. In addition, mutations of any of the cysteine residues of EGF in PE(ΔIa)-EGFmutants resulted in the loss of their binding activity to EGF receptor and a corresponding loss of their cytotoxicity. This study indicates that the cytotoxicity of PE(ΔIa)-EGFmutant to EGF receptor-bearing cells may be used as an indicator to screen mutations of EGF important in EGF-receptor interactions.

Original languageEnglish (US)
Pages (from-to)24034-24040
Number of pages7
JournalJournal of Biological Chemistry
Volume267
Issue number33
StatePublished - Nov 25 1992
Externally publishedYes

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Mutant Chimeric Proteins
Epidermal Growth Factor Receptor
Epidermal Growth Factor
Amino Acids
Cytotoxicity
Mutation
Bearings (structural)
Pseudomonas aeruginosa toxA protein
Immunotoxins
Mutagenesis
Point Mutation

ASJC Scopus subject areas

  • Biochemistry

Cite this

Pseudomonas exotoxin A-epidermal growth factor (EGF) mutant chimeric protein as an indicator for identifying amino acid residues important in EGF-receptor interaction. / Shiah, Her Shyong; Chen, Tzong Yueh; Chang, Chi Ming; Chow, Judy T.; Kung, Hsing-Jien; Hwang, Jaulang.

In: Journal of Biological Chemistry, Vol. 267, No. 33, 25.11.1992, p. 24034-24040.

Research output: Contribution to journalArticle

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