Characterization of poly-gamma-glutamate hydrolase encoded by a bacteriophage genome: possible role in phage infection of Bacillus subtilis encapsulated with poly-gamma-glutamate.
Some Bacillus subtilis strains, including natto (fermented soybeans) starter strains, produce a capsular polypeptide of glutamate with a gamma-linkage, called poly-gamma-glutamate (gamma-PGA). We identified and purified a monomeric 25-kDa degradation enzyme for gamma-PGA (designated gamma-PGA hydrolase, PghP) from bacteriophage PhiNIT1 in B. subtilis host cells. The monomeric PghP internally hydrolyzed gamma-PGA to oligopeptides, which were then specifically converted to tri-, tetra-, and penta-gamma-glutamates. Monoiodoacetate and EDTA both inhibited the PghP activity, but Zn(2+) or Mn(2+) ions fully restored the enzyme activity inhibited by the chelator, suggesting that a cysteine residue(s) and these metal ions participate in the catalytic mechanism of the enzyme. The corresponding pghP gene was cloned and sequenced from the phage genome. The deduced PghP sequence (208 amino acids) with a calculated M(r) of 22,939 was not significantly similar to any known enzyme. Thus, PghP is a novel gamma-glutamyl hydrolase. Whereas phage PhiNIT1 proliferated in B. subtilis cells encapsulated with gamma-PGA, phage BS5 lacking PghP did not survive well on such cells. Moreover, all nine phages that contaminated natto during fermentation produced PghP, supporting the notion that PghP is important in the infection of natto starters that produce gamma-PGA. Analogous to polysaccharide capsules, gamma-PGA appears to serve as a physical barrier to phage absorption. Phages break down the gamma-PGA barrier via PghP so that phage progenies can easily establish infection in encapsulated cells.
Kimura
Keitarou
K
Division of Applied Microbiology, National Food Research Institute, Kannondai 2-1-12, Tsukuba, Ibaraki 305-8642, Japan.
Itoh
Yoshifumi
Y
D000595:Amino Acid Sequence; D017103:Bacillus Phages / Q000201:enzymology* / Q000235:genetics* / Q000472:pathogenicity; D001412:Bacillus subtilis / Q000821:virology*; D001483:Base Sequence; D003001:Cloning, Molecular; D004279:DNA, Viral / Q000235:genetics; D016679:Genome, Viral; D008969:Molecular Sequence Data; D014774:Virulence; D011623:gamma-Glutamyl Hydrolase / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism
D004279:DNA, Viral; D011623:gamma-Glutamyl Hydrolase