Characterization of a haemolysin from Mycobacterium tuberculosis with homology to a virulence factor of Serpulina hyodysenteriae

• Published in: Microbiology (Society for General Microbiology) Vol. 144; no. 5; pp. 1205 - 1211
• Main Authors: Wren, B.W, Stabler, R.S, Das, S.S, Butcher, P.D, Mangan, J.A, Clarke, J.D, Casali, N, Parish, T, Stoker, N.G
• Format: Journal Article
• Language: English
• Published: Reading Soc General Microbiol 01.05.1998; Society for General Microbiology
• Subjects: Amino Acid Sequence; animal health; animal parasites and pests; Animals; arthropod pests; Bacterial Proteins; Bacteriology; Biological and medical sciences; Brachyspira hyodysenteriae - chemistry; Brachyspira hyodysenteriae - genetics; Brachyspira hyodysenteriae - pathogenicity; Cloning, Molecular; Fundamental and applied biological sciences. Psychology; genbank/x98295; Genes, Bacterial; Genetics; Hemolysin Proteins - chemistry; Hemolysin Proteins - genetics; Hemolysin Proteins - metabolism; Hemolysis; human health and safety; Humans; medicine; Microbiology; Molecular Sequence Data; Mycobacterium - chemistry; Mycobacterium - genetics; Mycobacterium leprae; Mycobacterium tuberculosis; Mycobacterium tuberculosis - chemistry; Mycobacterium tuberculosis - genetics; Polymerase Chain Reaction; Sequence Analysis, DNA; Sequence Homology, Amino Acid; Serpulina hyodysenteriae; Species Specificity; Transcription, Genetic; Virulence - genetics; Toxin; Mycobacterium tuberculosis; Nucleotide sequence; Mycobacteriales; Mycobacteriaceae; Bacteria; Homology; Actinomycetes; Gene expression; Hemolysin; Biological activity; Aminoacid sequence
• ISSN: 1350-0872; 1465-2080
• DOI: 10.1099/00221287-144-5-1205

Scrutiny of sequence data from the Mycobacterium leprae 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 Serpulina hyodysenteriae. Using degenerate oligonucleotide primers based on the TlyA sequences, the Mycobacterium tuberculosis homologue was amplified and this product was used to obtain the clone and sequence a 2.5 kb fragment containing the whole M. tuberculosis tlyA gene. tlyA encodes a 267-amino-acid protein with a predicted molecular mass of 28 kDa. TlyA homologues were identified by PCR in M. leprae, Mycobacterium avium and Mycobacterium bovis BCG, but appeared absent in Mycobacterium smegmatis, Mycobacterium vaccae, Mycobacterium kansasii, Mycobacterium chelonae and Mycobacterium phlei. The M. tuberculosis gene appeared to be the first gene in an operon containing at least two other genes. Introduction of the M. tuberculosis tlyA gene into M. smegmatis 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 tlyA was expressed as a His6-tagged fusion protein in Escherichia coli. tlyA mRNA was detected in both M. tuberculosis and M. bovis BCG using RT-PCR, confirming that this gene is expressed in organisms cultured in vitro.