Transposon mutagenesis and cloning of the genes encoding the enzymes of fengycin biosynthesis in Bacillus subtilis.
A total of 20 Bacillus subtilis F29-3 mutants defective in fengycin biosynthesis was obtained by Tn917 mutagenesis. Cloning and mapping results showed that the transposon in these mutants was inserted in eleven different locations on the chromosome. We were able to use the chromosomal sequence adjacent to the transposon as a probe to screen for cosmid clones containing the fengycin biosynthesis genes. One of the clones obtained, pFC660, was 46 kb long. Eight transposon insertion sites were mapped within this plasmid. Among the eleven different mutants analyzed, four mutants had Tn917 inserted in regions which encoded peptide sequences similar to part of gramicidin S synthetase, surfactin synthetase, and tyrocidine synthetase. Our results suggest that fengycin is synthesized nonribosomally by the multienzyme thiotemplate mechanism.
Chen
C L
CL
Institute of Botany, National Chung-Hsing University, Taichung, Taiwan.
Chang
L K
LK
Chang
Y S
YS
Liu
S T
ST
Tschen
J S
JS
D000251:Adenosine Triphosphatases / Q000235:genetics; D000591:Amino Acid Isomerases / Q000737:chemistry / Q000235:genetics; D000595:Amino Acid Sequence; D000935:Antifungal Agents / Q000096:biosynthesis*; D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics*; D001426:Bacterial Proteins; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D017124:Conserved Sequence / Q000235:genetics; D004251:DNA Transposable Elements / Q000235:genetics*; D055666:Lipopeptides; D008074:Lipoproteins / Q000096:biosynthesis / Q000235:genetics; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000737:chemistry / Q000235:genetics; D016254:Mutagenesis, Insertional; D010453:Peptide Synthases / Q000737:chemistry / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D014170:Transformation, Genetic
D000935:Antifungal Agents; D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D055666:Lipopeptides; D008074:Lipoproteins; D009097:Multienzyme Complexes; C049972:fengycin; D000251:Adenosine Triphosphatases; D000591:Amino Acid Isomerases; C022343:phenylalanine racemase (ATP-hydrolyzing); D010453:Peptide Synthases; C020831:gramicidin-S-synthetase 2; C069542:surfactin synthetase; C035180:tyrocidine synthetase