Sequence analysis and expression in Escherichia coli of the hyaluronidase gene of Streptococcus pyogenes bacteriophage H4489A.
The hyaluronidase gene (hylP) from Streptococcus pyogenes bacteriophage H4489A was previously cloned into Escherichia coli plasmid pUC8 as a 3.1-kilobase ThaI fragment. Southern hybridization experiments confirmed the origin of this fragment in bacteriophage H4489A before determination of the nucleotide sequence of the entire fragment. Two open reading frames (ORFs) were found, the first of which specified a 39,515-molecular-weight protein identified as the bacteriophage hyaluronidase. The second ORF encoded a 65,159-molecular-weight protein of unknown function. Putative transcription and translation control sequences for each ORF were identified by using a plasmid containing a promoterless chloramphenicol acetyltransferase gene. Controlled exclusive expression of the hylP gene via the T7 polymerase-promoter system in E. coli resulted in a 40,000-dalton protein, a result consistent with the coding capacity of the hylP gene.
Hynes
W L
WL
Department of Microbiology and Immunology, University of Oklahoma Health Sciences Center, Oklahoma City 73190.
Ferretti
J J
JJ
D000595:Amino Acid Sequence; D001435:Bacteriophages / Q000201:enzymology* / Q000235:genetics; D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D012321:DNA-Directed RNA Polymerases / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D005822:Genetic Vectors; D006821:Hyaluronoglucosaminidase / Q000235:genetics* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D011401:Promoter Regions, Genetic; D013297:Streptococcus pyogenes / Q000201:enzymology / Q000235:genetics*; D014764:Viral Proteins / Q000235:genetics / Q000302:isolation & purification
D014764:Viral Proteins; D012321:DNA-Directed RNA Polymerases; D006821:Hyaluronoglucosaminidase