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Infection and Immunity, July 1999, p. 3297-3301, Vol. 67, No. 7
0019-9567/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Differences in the Carboxy-Terminal (Putative Phospholipid Binding) Domains of Clostridium perfringens and Clostridium bifermentans Phospholipases C Influence the Hemolytic and Lethal Properties of These Enzymes

Marie Jepson,1,* Angela Howells,1 Helen L. Bullifent,1 Barbara Bolgiano,2 Dennis Crane,2 Julie Miller,1 Jane Holley,1 Pramukh Jayasekera,1 and Richard W. Titball1

Defence Evaluation and Research Agency, CBD Porton Down, Salisbury, Wiltshire SP4 0JQ,1 and National Institute for Biological Standards and Control, Potters Bar, Hertfordshire, EN6 3QG,2 United Kingdom

Received 23 December 1998/Returned for modification 18 February 1999/Accepted 3 April 1999

The phospholipases C of C. perfringens (alpha-toxin) and C. bifermentans (Cbp) show >50% amino acid homology but differ in their hemolytic and toxic properties. We report here the purification and characterisation of alpha-toxin and Cbp. The phospholipase C activity of alpha-toxin and Cbp was similar when tested with phosphatidylcholine in egg yolk or in liposomes. However, the hemolytic activity of alpha-toxin was more than 100-fold that of Cbp. To investigate whether differences in the carboxy-terminal domains of these proteins were responsible for differences in the hemolytic and toxic properties, a hybrid protein (NbiCalpha ) was constructed comprising the N domain of Cbp and the C domain of alpha-toxin. The hemolytic activity of NbiCalpha was 10-fold that of Cbp, and the hybrid enzyme was toxic. These results confirm that the C-terminal domain of these proteins confers different properties on the enzymatically active N-terminal domain of these proteins.

* Corresponding author. Mailing address: Defence Evaluation and Research Agency, CBD Porton Down, Salisbury, Wiltshire SP4 0JQ, United Kingdom. Phone: 1980-613301. Fax: 1980-613284. E-mail: 100655,2360{at}compuserve.com.

Infection and Immunity, July 1999, p. 3297-3301, Vol. 67, No. 7
0019-9567/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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