Scrutiny of sequence data from the genome sequencing project identified the presence of a gene encoding a 268-amino-acid polypeptide which is highly similar to a pore-forming haemolysin/cytotoxin virulence determinant, TlyA, from the swine pathogen Using degenerate oligonucleotide primers based on the TlyA sequences, the homologue was amplified and this product was used to obtain the clone and sequence a 2.5 kb fragment containing the whole gene. encodes a 267-amino-acid protein with a predicted molecular mass of 28 kDa. TlyA homologues were identified by PCR in and BCG, but appeared absent in and The gene appeared to be the first gene in an operon containing at least two other genes. Introduction of the gene into using a mycobacterial shuttle expression plasmid converted non-haemolytic cells into those exhibiting significant haemolytic activity. Similarly, inducible haemolytic activity was observed in sonicated bacteria when was expressed as a His-tagged fusion protein in mRNA was detected in both and BCG using RT-PCR, confirming that this gene is expressed in organisms cultured in


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