Characterization of rhamnolipids produced by wild-type and engineered Burkholderia kururiensis

Abstract

Biosurfactants are a class of functional molecules produced and secreted by microorganisms, which play important roles in cell physiology such as flagellum-dependent or -independent bacterial spreading, cell signaling, and biofilm formation. They are amphipathic compounds and comprise a variety of chemical structures, including rhamnolipids, typically produced by Pseudomonas spp. and also reported within other bacterial genera. The present study is focused on Burkholderia kururiensis KP23(T), a trichloroethylene (TCE)-degrading, N-fixing, and plant growth-promoting bacterium. Herein, we describe the production of rhamnolipids by B. kururiensis, and its characterization by LTQ-Orbitrap Hybrid Mass Spectrometry, a powerful tool that allowed efficient identification of molecular subpopulations, due to its high selectivity, mass accuracy, and resolving power. The population of rhamnolipids produced by B. kururiensis revealed molecular species commonly observed in Pseudomonas spp. and/or Burkholderia spp. In addition, this strain was used as a platform for expression of two Pseudomonas aeruginosa biosynthetic enzymes: RhlA, which directly utilizes β-hydroxydecanoyl-ACP intermediates in fatty acid synthesis to generate the HAA, and RhlB, the rhamnosyltransferase 1, which catalyzes the transfer of dTDP-L-rhamnose to β-hydroxy fatty acids in the biosynthesis of rhamnolipids. We show that rhamnolipid production by the engineered B. kururiensis was increased over 600 % when compared to the wild type. Structural analyses demonstrated a molecular population composed mainly of monorhamnolipids, as opposed to wild-type B. kururiensis and P. aeruginosa in which dirhamnolipids are predominant. We conclude that B. kururiensis is a promising biosurfactant-producing organism, with great potential for environmental and biotechnological applications due to its non-pathogenic characteristics and efficiency as a platform for metabolic engineering and production of tailor-made biosurfactants.

Dirhamnose-lipid production by recombinant nonpathogenic bacterium Pseudomonas chlororaphis

“…chlororaphis NRRL B-30761 is the first reported wildtype nonpathogenic organism that solely produces R 1 L (Gunther et al 2005(Gunther et al , 2007. Other reported RL-producing wild-type bacteria, including the non-Pseudomonas species (Hošková et al 2013;Řezanka et al 2011;Tavares et al 2013), synthesize a mixture of R 1 L and R 2 L. Therefore, in the present study, we genetically engineer the nonpathogenic NRRL B-30761 into an R 2 L-producing strain by expressing a heterologous rhlC gene in the organism. The resultant recombinant, P. chlororahis [pBS29-P2-rhlC], was shown to produce R 2 L at a level that is 2.4 times higher than R 1 L ( Table 2).…”

“…Rhamnolipids (RLs) are a family of rhamnose-containing glycolipids first characterized in Pseudomonas aeruginosa (Jarvis and Johnson 1949) and had since been described in other bacteria (Gunther et al 2005(Gunther et al , 2007Řezanka et al 2011;Tavares et al 2013;Toribio et al 2010). The lipid portion of most RLs consists of 3-hydroxyalkanoyl-3-hydroxyalkanoate (C x -C y , where x and y are the carbon chain lengths of the alkanoate) moiety, though some may contain only a monomeric 3-hydroxyalkanoate.…”

Production of Biosurfactants Using Eco-friendly Microorganisms

“…Pathogenicity of biosurfactant producers is key factor to prevent large-scale production; therefore, the nonpathogenic eco-friendly organisms need to be explored further, and a library of collections is important for the future development in this area of study. Tavares et al ( 2013 ) …”

“…Rhamnolipids produced by the engineered strains were mainly mono-RL as opposed to wild-type strains B. kururiensis and P. aeruginosa that predominantly produce di-RLs. This organism is a promising biosurfactant producer with potential environmental and biotechnological application especially due to its nonpathogenicity and compatibility with metabolic engineering procedures (Tavares et al 2013 ).…”

An Indigenous Biosurfactant Producing Burkholderia cepacia with High Emulsification Potential towards Crude Oil

“…In recent years, several studies have shown that a number of Burkholderia species such as Burkholderia thailandensis [34,35], Burkholderia glumae [36], Burkholderia kururiensis [37], Burkholderia plantarii [38], and Burkholderia gladioli [39] were able to produce biosurfactant. However, the ability of Burkholderia cepacia to produce a biosurfactant was not sufficiently documented in the literature.…”