Transposon mutagenesis was employed to isolate the gene(s) related with the
biosynthesis of dipeptide antibiotic in Bacillus subtilis PY79 (a prototro
phic derivative of the standard 168 strain). The blocked mutants were pheno
typically selected from the transposon library by bioassay and the complete
loss of biosynthetic ability was verified through ESI-mass spectrometry an
alysis. Four different bacilysin nonproducer mutants (Bac(-): :Tn10(ori-spc
)) were isolated from the transposon library. The genes involved in bacilys
in biosynthesis were identified as thyA (thymidilate synthetase), ybgG (unk
nown; similar to homocysteine methyl transferase) and oppA (oligopeptide pe
rmease), respectively. The other blocked gene was yvgW (unknown; similar to
heavy metal-transporting ATPase): however, backcross studies did not verif
y its involvement in bacilysin biosynthesis. This gene, on the other hand.
appeared to be necessary for efficient sporulation and transformation. Opp
involvement was significant as it suggested that bacilysin biosynthesis is
under or a component of the quorum sensing pathway which has been shown to
be responsible for the establishment of sporulation, competence development
and onset of surfactin biosynthesis. For verification, it was necessary to
check the involvement of peptide pheromones (PhrA or PhrC) internalized by
the Opp system and response regulator ComA as the essential components of
this global control. phrA, phrC and comA deleted mutants of PY79 were thus
constructed and the latter two genes were shown to be essential for bacilys
in biosynthesis. (C) 2001 Elsevier Science B.V. All rights reserved.