Mutagenesis of ISP5230 and selection for -aminobenzoic acid-dependent growth in the presence of sulfanilamide yielded mutants (VS519 and VS620) that continued to produce chloramphenicol (Cm), although with increased medium dependence. Transforming the mutants with pDQ102 or pDQ103, which carried a -complementing fragment from ISP5230 in alternative orientations, restored uniformly high Cm production in VS620, but did not alter the medium dependence of Cm production in VS519. The cloned DNA fragment was subcloned and trimmed to the minimum size conferring complementation. The resulting 2.8 kb HI-I fragment was sequenced. Codon preference analysis showed one complete ORF encoding a polypeptide of 670 amino acids. Comparison of the deduced amino acid sequence with database proteins indicated that the N- and C-terminal regions resembled PabA and PabB, respectively, of numerous bacteria. The gene product showed overall sequence similarity to the product of a fused gene associated with secondary metabolism in Insertion of an apramycin resistance gene into cloned in a segregationally unstable vector and replacement of the chromosomal with the disrupted copy lowered sulfanilamide resistance from 25 to 5 μg ml and blocked Cm production.


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