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Rhamnolipids are a class of biosurfactants which contain rhamnose as the sugar moiety linked to β-hydroxylated fatty acid chains. Rhamnolipids can be widely applied in many industries including petroleum, food, agriculture and bioremediation etc. Pseudomonas aeruginosa is still the most competent producer of rhamnolipids, but its pathogenicity may cause safety and health concerns during large-scale production and applications. Therefore, extensive studies have been carried out to explore safe and economical methods to produce rhamnolipids. Various metabolic engineering efforts have also been applied to either P. aeruginosa for improving its rhamnolipid production and diminishing its pathogenicity, or to other non-pathogenic strains by introducing the key genes for safe production of rhamnolipids. The three key enzymes for rhamnolipid biosynthesis, RhlA, RhlB and RhlC, are found almost exclusively in Pseudomonas sp. and Burkholderia sp., but have been successfully expressed in several non-pathogenic host bacteria to produce rhamnolipids in large scales. The composition of mono- and di-rhamnolipids can also be modified through altering the expression levels of RhlB and RhlC. In addition, cell-free rhamnolipid synthesis by using the key enzymes and precursors from non-pathogenic sources is thought to not only eliminate pathogenic effects and simplify the downstream purification processes, but also to circumvent the complexity of quorum sensing system that regulates rhamnolipid biosynthesis. The pathogenicity of P. aeruginosa can also be reduced or eliminated through in vivo or in vitro enzymatic degradation of the toxins such as pyocyanin during rhamnolipid production. The rhamnolipid production cost can also be significantly reduced if rhamnolipid purification step can be bypassed, such as utilizing the fermentation broth or the rhamnolipid-producing strains directly in the industrial applications of rhamnolipids.

Background: Oil palm empty fruit bunch (EFB) is a lignocellulosic waste produced in palm oil industry. EFB mainly consists of cellulose, hemicellulose (mainly xylan) and lignin and has a great potential to be reused. Converting EFB to fermentable sugars and value-added chemicals is a much better choice than treating EFB as waste., Results: A cellulase-producing strain growing on oil palm empty fruit bunch (EFB) was isolated and identified as Neurospora crassa S1, which is able to produce cellulases using EFB as the sole carbon source. The strain started to secret cellulases into the medium after 24 h of cultivation at 30°C and reached its maximal cellulase activity at 240 h. Mass spectroscopy (MS) analysis showed that more than 50 proteins were secreted into the medium when EFB was used as the sole carbon source. Among them, 7 proteins were identified as putative enzymes associated with cellulose degradation. The whole cell culture of Neurospora crassa S1 was used to hydrolyze acid-treated EFB, giving a total sugar yield of 83.2%, which is comparable with that (82.0%) using a well-known cellulase producer Trichoderma reesei RUT-C30 (ATCC56765)., Conclusion: Neurospora crassa S1 is a commercially promising native cellulase producer for EFB hydrolysis especially when the sugars obtained are to be fermented to products that require use of non-genetically engineered strains.

Background: Receptors with a single transmembrane (TM) domain are essential for the signal transduction across the cell membrane. NMR spectroscopy is a powerful tool to study structure of the single TM domain. The expression and purification of a TM domain in Escherichia coli (E.coli) is challenging due to its small molecular weight. Although ketosteroid isomerase (KSI) is a commonly used affinity tag for expression and purification of short peptides, KSI tag needs to be removed with the toxic reagent cyanogen bromide (CNBr)., Result: The purification of the TM domain of p75 neurotrophin receptor using a KSI tag with the introduction of a thrombin cleavage site is described herein. The recombinant fusion protein was refolded into micelles and was cleaved with thrombin. Studies showed that purified protein could be used for structural study using NMR spectroscopy., Conclusions: These results provide another strategy for obtaining a single TM domain for structural studies without using toxic chemical digestion or acid to remove the fusion tag. The purified TM domain of p75 neurotrophin receptor will be useful for structural studies.