Gene XV of bacteriophage PRD1 encodes a lytic enzyme with muramidase activity.
Bacteriophage PRD1 is a lipid-containing virus that infects a variety of Gram-negative bacteria, including Escherichia coli. The phage lyses its host by virtue of a virally-encoded lytic enzyme, the synthesis of which has been assigned to gene XV on the basis of complementation analysis and experiments with mutant phages. We report here the cloning of gene XV into an expression plasmid and the purification of its product, protein P15, to near homogeneity. The purified protein P15, identified by N-terminal sequence analysis, showed a strong lytic activity against chloroform-treated Gram-negative cells. No activity against Gram-positive bacterial species could be detected. The pH optimum of the enzyme was between 7.0-8.0. Protein P15 was readily inactivated at temperatures above 4 degrees C, as well as by increasing the ionic strength of the buffers. The analysis of cell wall digests indicated that P15 is a glycosidase that cleaves the beta (1-4) linkage between N-acetylmuramic acid and N-acetylglucosamine, thus displaying muramidase activity.
Caldentey
J
J
Institute of Biotechnology, University of Helsinki, Finland.
Hänninen
A L
AL
Bamford
D H
DH
D000595:Amino Acid Sequence; D001435:Bacteriophages / Q000201:enzymology* / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D004926:Escherichia coli / Q000821:virology; D015870:Gene Expression; D005814:Genes, Viral; D005816:Genetic Complementation Test; D006090:Gram-Negative Bacteria / Q000821:virology; D007700:Kinetics; D008969:Molecular Sequence Data; D009113:Muramidase / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D013045:Species Specificity; D014764:Viral Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism
D017931:DNA Primers; D011994:Recombinant Proteins; D014764:Viral Proteins; C090044:P15 protein, Enterobacteria phage PRD1; D009113:Muramidase