Identification of the carboxyl-terminal amino acids important for the ADP-ribosylation activity of Pseudomonas exotoxin A

J. T. Chow, M. S. Chen, H. C P Wu, J. Hwang

研究成果: 雜誌貢獻文章

15 引文 (Scopus)

摘要

The ADP-ribosylation domain of Pseudomonas exotoxin A (PE) has been identified to reside in structural domain III (residues 405-613) and a portion of domain Ib (residues 385-404) of the molecule (Hwang, J., FitzGerald, D.J., Adhya, S., and Pastan, I., (1987) Cell 48, 128-136). To further determine the carboxyl end region essential for ADP-ribosylation activity, we constructed sequential deletions at the carboxyl-terminal of PE. Our results show that a clone with a deletion of the carboxyl-terminal amino acid residues from Arg-609 to Lys-613 and replaced with Arg-Asn retained wild-type PE ADP-ribosylation activity. Deletion of the terminal amino acid residues from Ala-596 to Lys-613 and replaced with Val-Ile-Asn reduced ADP-ribosylation activity by 75%, while deletions of 36 or more amino acids from the carboxyl terminus completely lose their ADP-ribosylation activity. These modified PEs were also examined for their ability to block PE cytotoxicity. Our results show that modified PEs which lost their ADP-ribosylation activity correspondingly lost their cytotoxicity. Furthermore, extracts containing PE fragments without ADP-ribosylation activty were able to block the cytotoxic activity of intact PE. Our results thus indicate that carboxyl-terminal amino acids in the Ser-595 region are crucial for ADP-ribosylation activity and, consequently, cytotoxicity of PE. The modified PEs which have lost their ADP-ribosylation activity may also be a route to new PE vaccines.

原文英語
頁(從 - 到)18818-18823
頁數6
期刊Journal of Biological Chemistry
264
發行號31
出版狀態已發佈 - 1989
對外發佈Yes

指紋

Adenosine Diphosphate
Amino Acids
Cytotoxicity
Pseudomonas aeruginosa toxA protein
Vaccines
Clone Cells
Molecules

ASJC Scopus subject areas

  • Biochemistry

引用此文

Identification of the carboxyl-terminal amino acids important for the ADP-ribosylation activity of Pseudomonas exotoxin A. / Chow, J. T.; Chen, M. S.; Wu, H. C P; Hwang, J.

於: Journal of Biological Chemistry, 卷 264, 編號 31, 1989, p. 18818-18823.

研究成果: 雜誌貢獻文章

Chow, J. T. ; Chen, M. S. ; Wu, H. C P ; Hwang, J. / Identification of the carboxyl-terminal amino acids important for the ADP-ribosylation activity of Pseudomonas exotoxin A. 於: Journal of Biological Chemistry. 1989 ; 卷 264, 編號 31. 頁 18818-18823.
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abstract = "The ADP-ribosylation domain of Pseudomonas exotoxin A (PE) has been identified to reside in structural domain III (residues 405-613) and a portion of domain Ib (residues 385-404) of the molecule (Hwang, J., FitzGerald, D.J., Adhya, S., and Pastan, I., (1987) Cell 48, 128-136). To further determine the carboxyl end region essential for ADP-ribosylation activity, we constructed sequential deletions at the carboxyl-terminal of PE. Our results show that a clone with a deletion of the carboxyl-terminal amino acid residues from Arg-609 to Lys-613 and replaced with Arg-Asn retained wild-type PE ADP-ribosylation activity. Deletion of the terminal amino acid residues from Ala-596 to Lys-613 and replaced with Val-Ile-Asn reduced ADP-ribosylation activity by 75{\%}, while deletions of 36 or more amino acids from the carboxyl terminus completely lose their ADP-ribosylation activity. These modified PEs were also examined for their ability to block PE cytotoxicity. Our results show that modified PEs which lost their ADP-ribosylation activity correspondingly lost their cytotoxicity. Furthermore, extracts containing PE fragments without ADP-ribosylation activty were able to block the cytotoxic activity of intact PE. Our results thus indicate that carboxyl-terminal amino acids in the Ser-595 region are crucial for ADP-ribosylation activity and, consequently, cytotoxicity of PE. The modified PEs which have lost their ADP-ribosylation activity may also be a route to new PE vaccines.",
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