T al. AMB Express 2013, three:66 amb-express/content/3/1/ORIGINAL ARTICLEOpen AccessOptimisation of engineered Escherichia coli biofilms for

T al. AMB Express 2013, three:66 amb-express/content/3/1/ORIGINAL ARTICLEOpen AccessOptimisation of engineered Escherichia coli biofilms for enzymatic biosynthesis of L-halotryptophansStefano Perni1, Louise Hackett1, Rebecca JM Goss2, Mark J Simmons1 and Tim W Overton1AbstractEngineered biofilms comprising a single recombinant PKCη Accession species have demonstrated remarkable activity as novel biocatalysts for a array of applications. In this function, we focused around the biotransformation of 5-haloindole into 5-halotryptophan, a pharmaceutical intermediate, making use of Escherichia coli expressing a recombinant tryptophan synthase enzyme encoded by plasmid pSTB7. To optimise the reaction we compared two E. coli K-12 strains (MC4100 and MG1655) and their ompR234 mutants, which overproduce the adhesin curli (PHL644 and PHL628). The ompR234 mutation elevated the quantity of biofilm in each MG1655 and MC4100 backgrounds. In all circumstances, no conversion of 5-haloindoles was observed utilizing cells with no the pSTB7 plasmid. Engineered biofilms of strains PHL628 pSTB7 and PHL644 pSTB7 generated more 5-halotryptophan than their corresponding planktonic cells. Flow cytometry revealed that the vast majority of cells were alive soon after 24 hour biotransformation reactions, each in planktonic and biofilm types, suggesting that cell viability was not a significant issue within the higher overall performance of biofilm reactions. Monitoring 5-haloindole depletion, 5-halotryptophan synthesis plus the percentage conversion from the biotransformation reaction recommended that there had been inherent variations among strains MG1655 and MC4100, and involving planktonic and biofilm cells, when it comes to tryptophan and indole metabolism and transport. The study has reinforced the will need to completely investigate bacterial physiology and make informed strain selections when establishing biotransformation reactions. Keywords: E. coli; Biofilm; Biotransformation; Haloindole; HalotryptophanIntroduction Bacterial biofilms are renowned for their enhanced resistance to environmental and chemical stresses including antibiotics, metal ions and organic solvents when in comparison with planktonic bacteria. This house of biofilms is often a cause of clinical concern, specifically with implantable healthcare devices (which include catheters), considering the fact that biofilm-mediated infections are often tougher to treat than those caused by planktonic bacteria (Smith and Hunter, 2008). However, the improved robustness of biofilms might be exploited in bioprocesses exactly where cells are exposed to harsh reaction circumstances (Winn et al., 2012). Biofilms, normally multispecies, happen to be employed for waste water therapy (biofilters) (Purswani et al., 2011; Iwamoto and Nasu, 2001; Correspondence: [email protected] 1 College of Chemical Engineering, University of Birmingham, Birmingham B15 2TT, UK Full list of author details is readily available in the finish in the articleCortes-Lorenzo et al., 2012), air filters (Rene et al., 2009) and in soil bioremediation (Zhang et al., 1995; Singh and Cameotra, 2004). Most not too long ago, single species biofilms have discovered applications in microbial fuel cells (Yuan et al., 2011a; Yuan et al., 2011b) and for specific biocatalytic reactions (Tsoligkas et al., 2011; Gross et al., 2010; Kunduru and Pometto, 1996). Recent examples of biotransformations catalysed by single-species biofilms include things like the conversion of benzaldehyde to benzyl alcohol (Zymomonas mobilis; Li et al., 2006), ethanol production (Z. Epoxide Hydrolase Inhibitor Formulation mobilis and Saccharomyces cerevisiae; Kunduru and Pomett.