Enhanced biosurfactant production through cloning of three genes and role of esterase in biosurfactant release

Abstract

BackgroundBiosurfactants have been reported to utilize a number of immiscible substrates and thereby facilitate the biodegradation of panoply of polyaromatic hydrocarbons. Olive oil is one such carbon source which has been explored by many researchers. However, studying the concomitant production of biosurfactant and esterase enzyme in the presence of olive oil in the Bacillus species and its recombinants is a relatively novel approach.ResultsBacillus species isolated from endosulfan sprayed cashew plantation soil was cultivated on a number of hydrophobic substrates. Olive oil was found to be the best inducer of biosurfactant activity. The protein associated with the release of the biosurfactant was found to be an esterase. There was a twofold increase in the biosurfactant and esterase activities after the successful cloning of the biosurfactant genes from Bacillus subtilis SK320 into E.coli. Multiple sequence alignment showed regions of similarity and conserved sequences between biosurfactant and esterase genes, further confirming the symbiotic correlation between the two. Biosurfactants produced by Bacillus subtilis SK320 and recombinant strains BioS a, BioS b, BioS c were found to be effective emulsifiers, reducing the surface tension of water from 72 dynes/cm to as low as 30.7 dynes/cm.ConclusionThe attributes of enhanced biosurfactant and esterase production by hyper-producing recombinant strains have many utilities from industrial viewpoint. This study for the first time has shown a possible association between biosurfactant production and esterase activity in any Bacillus species. Biosurfactant-esterase complex has been found to have powerful emulsification properties, which shows promising bioremediation, hydrocarbon biodegradation and pharmaceutical applications.

Biosurfactant Production and Potential Correlation with Esterase Activity

“…The production of surfactin in culture broth of Bacillus subtilis [16], rhamnolipids by Pseudomonas aeruginosa [17], exopolysaccharides in A. calcoaceticus BD4 [18] and rhamnolipid AP-6 in P. fluorescens 378 [19] were all found to be growth associated. In Bacillus subtilis SK320, the production of biosurfactant was also found to be growth associated [3]. Maximum biosurfactant activity was associated with cells at the stationary phase of growth when the nutrient limiting conditions started establishing in the culture medium.…”

“…The total yield reported as g/L was calculated after purifying the biosurfactant using acetone and weighing the lyophilized powder (Figure 1). The recombinant strains not only showed enhanced biosurfactant production, but an increase in the esterase activity as well as surface tension values (Table 4) [3]. Similarly, Bacillus clausii 5B when grown on minimal medium containing 1% (w/v) glucose as carbon source gave the total yield of 2.11 g/L after 96 h [36], whereas Kocuria turfanesis BS-J and Pseudomonas aeruginosa BS-P when grown on combined distillery waste with other industrial wastes showed the biosurfactant production of 0.967 g/L and 1.976 g/L, respectively [37].…”

“…Highly insoluble carbon source such as n-hexadecane, paraffinic oil, glycerol, babassu oil for P. aeruginosa PA1 [28], soybean curd residue (okara) for B. subtilis YB8 and B. subtilis MI113 [29,30], peat hydrolysate for B. subtilis [31], soybean oil, safflower oil, glycerol for P. aeruginosa GS9-119 and DS10-129 [32] and sludge palm oil for Klebsiella pneumoniae WMF02 [33] have been reported to induce high biosurfactant production. Olive oil was an unconventional carbon source used for the production of lipopeptide biosurfactant by Bacillus subtilis SK320 [3] and biosurfactant produced by marine Nocardiopsis B4 [34]. The important finding was the temperature stability of both the biosurfactants increasing their scope of application at higher temperatures like in microbial enhanced oil recovery.…”

“…Biosurfactants and esterases thus show some sort of symbiotic relationship which is mutually beneficial to them. Biosurfactants, produced by the recombinant strains, are found to be responsible for the enhanced production and activity of esterases and vice versa [3]. This can further be explained by the fact that biosurfactants are known to be poteinaceous in nature [4,5] and it is the protein moiety which is responsible for the emulsifying activity or detergent like property of biosurfactants.…”

Microbiotechnology Based Surfactants and Their Applications

“…The operating costs can be brought down by robust wild-type strains or recombinant mutant strains. Testing the possibility of co-products and/or enzymes is another attractive solution to surge the net profitfor example: esterases released during the production of lipopeptides by Bacillus strain and its recombinants (Sekhon et al, 2011. Co-products and by-products are value drivers and increase the economic viability of any business.…”