The bacitracin biosynthesis operon of Bacillus licheniformis ATCC 10716: molecular characterization of three multi-modular peptide synthetases.
BACKGROUND
The branched cyclic dodecylpeptide antibiotic bacitracin, produced by special strains of Bacillus, is synthesized nonribosomally by a large multienzyme complex composed of the three bacitracin synthetases BA1, BA2 and BA3. These enzymes activate and incorporate the constituent amino acids of bacitracin by a thiotemplate mechanism in a pathway driven by a protein template. The biochemical features of these enzymes have been studied intensively but little is known about the molecular organization of their genes.


RESULTS
The entire bacitracin synthetase operon containing the genes bacA-bacC was cloned and sequenced, identifying a modular structure typical of peptide synthetases. The bacA gene product (BA1, 598kDa) contains five modules, with an internal epimerization domain attached to the fourth; bacB encodes BA2 (297kDa), and has two modules and a carboxy-terminal epimerization domain; bacC encodes BA3, five modules (723kDa) with additional internal epimerization domains attached to the second and fourth. A carboxy-terminal putative thioesterase domain was also detected in BA3. A putative cyclization domain was found in BA1 that may be involved in thiazoline ring formation. The adenylation/thioester-binding domains of the first two BA1 modules were overproduced and the detected amino-acid specificity coincides with the first two amino acids in bacitracin. Disruption of chromosomal bacB resulted in a bacitracin-deficient mutant.


CONCLUSIONS
The genes encoding the bacitracin synthetases BA1, BA2 and BA3 are organized in an operon, the structure of which reflects the modular architecture expected of peptide synthetases. In addition, a putative thiazoline ring formation domain was identified in the BA1 gene.
Konz
D
D
Philipps-Universität Marburg Fachbereich Chemie/Biochemie Hans-Meerwein-Strasse, 35032, Marburg, Germany.
Klens
A
A
Schörgendorfer
K
K
Marahiel
M A
MA
D000595:Amino Acid Sequence; D001407:Bacillus / Q000235:genetics* / Q000378:metabolism*; D001414:Bacitracin / Q000096:biosynthesis*; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000032:analysis / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D009876:Operon / Q000235:genetics; D010453:Peptide Synthases / Q000235:genetics* / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D017262:Siderophores / Q000378:metabolism
D004269:DNA, Bacterial; D017262:Siderophores; D001414:Bacitracin; D010453:Peptide Synthases