Analysis of a second bacteriophage hyaluronidase gene from Streptococcus pyogenes: evidence for a third hyaluronidase involved in extracellular enzymatic activity.
The hyaluronidase gene (hylP2) from a second group A streptococcal bacteriophage was isolated from ATCC T-type-22 hyaluronidase-producing strain 10403, a strain known to produce increased amounts of extracellular hyaluronidase. Sequence analysis of hylP2 and alignment with the previously described bacteriophage hyaluronidase gene (hylP) showed a high degree of similarity; however, hylP2 had deletions of regions specifying 34 amino acids. Twenty-eight of the deleted amino acids were in a region of HylP containing a series of collagen-like Gly-X-Y repeating units. By employing primers for both hylP and hylP2, PCR amplification resulted in fragments of appropriate sizes in 97% of the strains tested, with some strains producing two fragments, indicating the presence of at least two phages. When the hylP2 gene was introduced via a plasmid vector into a non-hyaluronidase-producing Streptococcus pyogenes strain, this strain was still unable to produce extracellular hyaluronidase, although intracellular hyaluronidase was present. These results, along with the absence of a typical N-terminal signal peptide, indicate that HylP2 is unable to be secreted into the extracellular milieu. Examination of more than 100 strains for production of hyaluronidase showed that only 23% of the strains produced extracellular hyaluronidase. One of these strains (strain 10403) contains a single bacteriophage hyaluronidase gene (hylP2) which, when inactivated by allelic replacement, still produces large amounts of extracellular hyaluronidase. These results suggest the presence of a different hyaluronidase gene encoding a protein that is actively secreted into the extracellular milieu.
Hynes
W L
WL
Department of Microbiology and Immunology, University of Oklahoma Health Sciences Center, Oklahoma City 73190, USA.
Hancock
L
L
Ferretti
J J
JJ
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D005798:Genes, Bacterial; D006821:Hyaluronoglucosaminidase / Q000235:genetics*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D013297:Streptococcus pyogenes / Q000201:enzymology* / Q000235:genetics
D017931:DNA Primers; D006821:Hyaluronoglucosaminidase