The pobA gene of Burkholderia cenocepacia encodes a Group I Sfp-type phosphopantetheinyltransferase required for biosynthesis of the siderophores ornibactin and pyochelin

Abstract

The opportunistic pathogen Burkholderia cenocepacia produces the siderophores ornibactin and pyochelin under iron-restricted conditions. Biosynthesis of both siderophores requires the involvement of non-ribosomal peptide synthetases (NRPSs). Using a transposon containing the lacZ reporter gene, two B. cenocepacia mutants were isolated which were deficient in siderophore production. Mutant IW10 was shown to produce normal amounts of ornibactin but only trace amounts of pyochelin, whereas synthesis of both siderophores was abolished in AHA27. Growth of AHA27, but not IW10, was inhibited under iron-restricted conditions. In both mutants, the transposon had integrated into the pobA gene, which encodes a polypeptide exhibiting similarity to the Sfp-type phosphopantetheinyltransferases (PPTases). These enzymes are responsible for activation of NRPSs by the covalent attachment of the 4′-phosphopantetheine (P-pant) moiety of coenzyme A. Previously characterized PPTase genes from other bacteria were shown to efficiently complement both mutants for siderophore production when provided in trans. The B. cenocepacia pobA gene was also able to efficiently complement an Escherichia coli entD mutant for production of the siderophore enterobactin. Using mutant IW10, in which the lacZ gene carried by the transposon is inserted in the same orientation as pobA, it was shown that pobA is not appreciably iron-regulated. Finally, we confirmed that Sfp-type bacterial PPTases can be subdivided into two distinct groups, and we present the amino acid signature sequences which characterize each of these groups.

Burkholderia paludis sp. nov., an Antibiotic-Siderophore Producing Novel Burkholderia cepacia Complex Species, Isolated from Malaysian Tropical Peat Swamp Soil

“…The genus Burkholderia is well known for producing a wide range of secondary metabolites which include siderophores (Asghar et al, 2011), antifungal (Lu et al, 2009; Tawfik et al, 2010), antibacterial (El-Banna and Winkelmann, 1998; Mitchell and Teh, 2005; Mitchell et al, 2008), antitumor (Klausmeyer et al, 2011; He et al, 2014), and exotoxins (Jeong et al, 2003; Partida-Martinez and Hertweck, 2007). Secondary metabolites are compounds which are non-essential for microbial growth or reproduction but provide diverse survival functions in nature (Martin et al, 2005).…”

Investigation of the multifaceted iron acquisition strategies of Burkholderia cenocepacia

“…The patient was also infected with P. aeruginosa for the period during which the B. cenocepacia strains were isolated. Burkholderia cenocepacia 715j and the 715j pobA mutant, AHA27 ('715j-pobA'), have been described previously (Darling et al 1998;Asghar et al 2011). The pobA gene encodes a polypeptide exhibiting similarity to the Sfp-type phosphopantetheinyl transferases (PPTases) and is required for activation of the non-ribosomal peptide synthetases (NRPSs) OrbI, OrbJ, PchE and PchF required for ornibactin and pyochelin production, and as such 715j-pobA is unable to produce ornibactin or pyochelin (Asghar et al 2011).…”

“…Burkholderia cenocepacia 715j and the 715j pobA mutant, AHA27 ('715j-pobA'), have been described previously (Darling et al 1998;Asghar et al 2011). The pobA gene encodes a polypeptide exhibiting similarity to the Sfp-type phosphopantetheinyl transferases (PPTases) and is required for activation of the non-ribosomal peptide synthetases (NRPSs) OrbI, OrbJ, PchE and PchF required for ornibactin and pyochelin production, and as such 715j-pobA is unable to produce ornibactin or pyochelin (Asghar et al 2011). 715j-orbI was constructed by transfer of the orbI::mini-Tn5Tp allele from the B. cenocepacia ornibactin mutant OM13 (Agnoli et al 2006) using the allelic replacement vector pSHAFT2 (S. Shastri, K. Karan, V. A. Eidsvaag and M. S. Thomas, unpublished results) and was shown to produce pyochelin and not ornibactin by HPLC (data not shown).…”

Iron acquisition in the cystic fibrosis lung and potential for novel therapeutic strategies

“…To date, species known to produce ornibactin under iron starvation include Burkholderia cepacia, Burkholderia cenocepacia, Burkholderia vietnamiensis and Burkholderia ambifaria (Meyer et al, 1995;Thomas, 2007;Asghar et al, 2011). More recently, ornibactin gene expression has been reported for the first time in Burkholderia multivorans (Denman et al, 2014).…”