Nucleotide sequence of the hemolysin I gene from Actinobacillus pleuropneumoniae.
The DNA sequence of the gene encoding the structural protein of hemolysin I (HlyI) of Actinobacillus pleuropneumoniae serotype 1 strain 4074 was analyzed. The nucleotide sequence shows a 3,072-bp reading frame encoding a protein of 1,023 amino acids with a calculated molecular size of 110.1 kDa. This corresponds to the HlyI protein, which has an apparent molecular size on sodium dodecyl sulfate gels of 105 kDa. The structure of the protein derived from the DNA sequence shows three hydrophobic regions in the N-terminal part of the protein, 13 glycine-rich domains in the second half of the protein, and a hydrophilic C-terminal area, all of which are typical of the cytotoxins of the RTX (repeats in the structural toxin) toxin family. The derived amino acid sequence of HlyI shows 42% homology with the hemolysin of A. pleuropneumoniae serotype 5, 41% homology with the leukotoxin of Pasteurella haemolytica, and 56% homology with the Escherichia coli alpha-hemolysin. The 13 glycine-rich repeats and three hydrophobic areas of the HlyI sequence show more similarity to the E. coli alpha-hemolysin than to either the A. pleuropneumoniae serotype 5 hemolysin or the leukotoxin (while the last two are more similar to each other). Two types of RTX hemolysins therefore seem to be present in A. pleuropneumoniae, one (HlyI) resembling the alpha-hemolysin and a second more closely related to the leukotoxin. Ca(2+)-binding experiments using HlyI and recombinant A. pleuropneumoniae prohemolysin (HlyIA) that was produced in E. coli shows that HlyI binds 45Ca2+, probably because of the 13 glycine-rich repeated domains. Activation of the prohemolysin is not required for Ca2+ binding.
Frey
J
J
Institute for Veterinary Bacteriology, University of Berne, Switzerland.
Meier
R
R
Gygi
D
D
Nicolet
J
J
D000188:Actinobacillus / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D001427:Bacterial Toxins; D001483:Base Sequence; D002118:Calcium / Q000378:metabolism; D004269:DNA, Bacterial / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial / Q000235:genetics*; D006460:Hemolysin Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D016364:Reading Frames; D012689:Sequence Homology, Nucleic Acid
D001427:Bacterial Toxins; D004269:DNA, Bacterial; D006460:Hemolysin Proteins; D002118:Calcium