The Riddle of Rhamnolipid Molecular Biosynthesis

Rhamnolipids (RL) were first shown to be produced by the opportunistic pathogen Pseudomonas aeruginosa. RL production is achieved via a biosynthesis pathway comprising three separate enzymes; RhlA, responsible for the synthesis of the fatty acid dimer precursor moieties (HAA); RhlB, a rhamnosyltransferase enzyme which conjugates HAA to dTDP-L-rhamnose to form mono-RL and RhlC, a second rhamnosyltransferase enzyme that adds a second rhamnose to form di-RL. Within P. aeruginosa the genes encoding the first two enzymes, rhlA and rhlB, are present in a single operon alongside genes encoding an AHL-mediated quorum sensing system rhlR and rhlI. The gene encoding the last enzyme, rhlC, is located approx. 1 Mb downstream of this operon. Since the discovery of RL synthesis by P. aeruginosa other bacterial species have also been shown to synthesise RLs, notably Burkholderia thailandensis and Burkholderia pseudomallei. Interestingly the RL biosynthesis genes within Burkholderia only show 40% similarity to those of P. aeruginosa, are located together in a single operon and are duplicated within the genome to give two functional copies of each gene. Recently we have shown RL synthesis in two marine bacteria; Pseudomonas sp. MCTG214(3b1) and Marinobacter sp. MCTG107b. In the Pseudomonas sp. we identified rhlA and rhlB homologues with 99% similarity to those of P. aeruginosa, however we have yet to identify any rhlC homologue. This presents a paradox as we have demonstrated di-RL synthesis by this strain. RL synthesis within the Marinobacter sp. presents an even greater paradox as we have yet to identify any RL synthase homologs. Here we explain the molecular differences in RL synthase homologues, the effects that these differences may create and try to explain the apparent lack of synthesis genes in the two marine strains. We also discus the methods used to investigate the molecular mechanisms of RL synthesis.