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.
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.
Jepson
M
M
Defence Evaluation and Research Agency, CBD Porton Down, Salisbury, Wiltshire SP4 0JQ, United Kingdom.
Howells
A
A
Bullifent
H L
HL
Bolgiano
B
B
Crane
D
D
Miller
J
J
Holley
J
J
Jayasekera
P
P
Titball
R W
RW
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism / Q000633:toxicity; D003001:Cloning, Molecular; D003013:Clostridium / Q000201:enzymology*; D003015:Clostridium Infections / Q000201:enzymology / Q000382:microbiology*; D003016:Clostridium perfringens / Q000201:enzymology*; D004789:Enzyme Activation; D051379:Mice; D008969:Molecular Sequence Data; D010743:Phospholipids / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity; D010738:Type C Phospholipases / Q000235:genetics / Q000378:metabolism / Q000633:toxicity*
D001426:Bacterial Proteins; D010743:Phospholipids; D010738:Type C Phospholipases