p-Aminobenzoic acid and chloramphenicol biosynthesis in Streptomyces venezuelae: gene sets for a key enzyme, 4-amino-4-deoxychorismate synthase The GenBank accession number for the sequence reported in this paper is AF189258.

Abstract

Amplification of sequences from Streptomyces venezuelae ISP5230 genomic DNA using PCR with primers based on conserved prokaryotic pabB sequences gave two main products. One matched pabAB, a locus previously identified in S. venezuelae. The second closely resembled the conserved pabB sequence consensus and hybridized with a 3 8 kb NcoI fragment of S. venezuelae ISP5230 genomic DNA. Cloning and sequence analysis of the 3 8 kb fragment detected three ORFs, and their deduced amino acid sequences were used in BLAST searches of the GenBank database. The ORF1 product was similar to PabB in other bacteria and to the PabB domain encoded by S. venezuelae pabAB. The ORF2 product resembled PabA of other bacteria. ORF3 was incomplete ; its deduced partial amino acid sequence placed it in the MocR group of GntR-type transcriptional regulators. Introducing vectors containing the 3 8 kb NcoI fragment of S. venezuelae DNA into pabA and pabB mutants of Escherichia coli, or into the Streptomyces lividans pab mutant JG10, enhanced sulfanilamide resistance in the host strains. The increased resistance was attributed to expression of the pair of discrete translationally coupled p-aminobenzoic acid biosynthesis genes (designated pabB/pabA) cloned in the 3 8 kb fragment. These represent a second set of genes encoding 4-amino-4-deoxychorismate synthase in S. venezuelae ISP5230. In contrast to the fused pabAB set previously isolated from this species, they do not participate in chloramphenicol biosynthesis, but like pabAB they can be disrupted without affecting growth on minimal medium. The gene disruption results suggest that S. venezuelae may have a third set of genes encoding PABA synthase.

Biosynthesis of the dichloroacetyl component of chloramphenicol in Streptomyces venezuelae ISP5230: genes required for halogenation

“…Twelve of them are clustered on the approximately 18 kb of chromosomal DNA sequenced in this and previous studies (Mosher et al, 1990(Mosher et al, , 1995Brown et al, 1996;Chang et al, 2001;He et al, 2001). Functions for nine genes have been confirmed by insertional inactivation, and particular interest has focused on one (cmlP; see Fig.…”

“…S. venezuelae genes initially cloned in pUC18 or pBluescript II (SK+) were recloned in the conjugal E. coli plasmid pJV326 before insertion of the 1?5 kb apramycin resistance (Am R ) cassette from pUC120A (Paradkar & Jensen, 1995) or pJV225 (Chang et al, 2001) at a site near the centre of the cloned fragment. To disrupt cmlK (ORF11), conjugal plasmid pJV526 was linearized with NotI and ligated with Am R from pJV225 to give pJV527.…”

Biosynthesis of the dideoxysugar component of jadomycin B: genes in the jad cluster of Streptomyces venezuelae ISP5230 for l-digitoxose assembly and transfer to the angucycline aglycone The GenBank accession number for the sequence reported in this paper is AY026363.

“…The Am R gene was excised from pJV225 (Chang et al, 2001) as a HindIII fragment and ligated into the HindIII site of pXE4 (Ingram et al, 1989) with the same transcriptional orientation as xylE. The pXE4 : : Am R construct and a pXE4 control plasmid were passaged through E. coli ET12567 before being used to transform protoplasts of Streptomyces venezuelae ISP5230.…”

Characterization of the Amicetin Biosynthesis Gene Cluster from Streptomyces vinaceusdrappus NRRL 2363 Implicates Two Alternative Strategies for Amide Bond Formation

“…PABA has been shown to serve as the starter unit initiating the biosynthesis of aureothin (32) and FR-008/candicidin (7) and contributes to the skeleton of chloramphenicol (3). PABA moieties in these secondary metabolites are derived from chorismate via two enzymatic steps catalyzed by a PABA synthase and a 4-amino-4-deoxychorismate (ADC) lyase (6,32,61). The primary amino group of cytosine is linked by PABA, to which a (ϩ)-␣-methylserine residue is attached, forming two amide bonds in the amicetin structure.…”