A simple and scalable one-pot biotransformation enables direct access to l-halotryptophans, including l-7-iodotryptophan, from the corresponding haloindoles. The biotransformation utilizes an easy to prepare bacterial cell lysate that may be stored as the lyophilizate for several months and utilized as a catalyst as and when required.
Smith, DRM, Willemse T, Gkotsi DS, Schepens W, Maes BU, Ballet S, Goss RJM
Org Lett 2014, 16 (10) 2622-2625
Natural Products: Discourse, Diversity and Design provides an informative and accessible overview of discoveries in the area of natural products in the genomic era, bringing together advances across the kingdoms. As genomics data makes it increasingly clear that the genomes of microbes and plants contain far more genes for natural product synthesis than had been predicted from the numbers of previously identified metabolites, the potential of these organisms to synthesize diverse natural products is likely to be far greater than previously envisaged. Natural Products addresses not only the philosophical questions of the natural role of these metabolites, but also the evolution of single and multiple pathways, and how these pathways and products may be harnessed to aid discovery of new bioactives and modes of action.
Edited by recognized leaders in the fields of plant and microbial biology, bioorganic chemistry and natural products chemistry, and with contributions from researchers at top labs around the world, Natural Products is unprecedented in its combination of disciplines and the breadth of its coverage. Natural Produces: Discourse, Diversity and Design will appeal to advanced students and experienced researchers, from academia to industry, in diverse areas including ecology, industrial biotechnology, drug discovery, medicinal chemistry, agronomy, crop improvement, and natural product chemistry.
Osbourn A (ed), Goss R (ed), Carter GT (ed)
April 2014, Wiley-Blackwell. ISBN: 978-1-118-29806-0
Engineered biofilms comprising a single recombinant species have demonstrated remarkable activity as novel biocatalysts for a range of applications. In this work, we focused on the biotransformation of 5-haloindole into 5-halotryptophan, a pharmaceutical intermediate, using 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 increased the quantity of biofilm in both MG1655 and MC4100 backgrounds. In all cases, no conversion of 5-haloindoles was observed using cells without 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 after 24 hour biotransformation reactions, both in planktonic and biofilm forms, suggesting that cell viability was not a major factor in the greater performance of biofilm reactions. Monitoring 5-haloindole depletion, 5-halotryptophan synthesis and the percentage conversion of the biotransformation reaction suggested that there were inherent differences between strains MG1655 and MC4100, and between planktonic and biofilm cells, in terms of tryptophan and indole metabolism and transport. The study has reinforced the need to thoroughly investigate bacterial physiology and make informed strain selections when developing biotransformation reactions.
Perni S, Hackett L, Goss RJM, Simmons MJ, Overton TW
AMB Express 2013, 3:66
Smith DRM; Grüschow S; Goss RJM
Curr Opin Chem Biol 2012, 17 (2) 276-283
A large and diverse series of halogenated natural products exist. In many of these compounds the halogen is important to biological activity and bioavailability. We now recognise that nature has developed many different halogenation strategies for which well-known enzyme classes such as haem oxidases or flavin-dependent oxidases have been adapted. Enzymes capable of halogenating all kinds of different chemical groups from electron-rich to electron-poor, from aromatic to aliphatic have been characterised. Given that synthetic halogenation reactions are not trivial transformations and that halogenated molecules possess pharmaceutical usefulness, it will be worth investing into further research of halogenating enzymes.
Amorn Owatworakit ; Belinda Townsend ; Thomas Louveau ; Helen Jenner ; Martin Rejzek ; Richard K. Hughes ; Gerhard Saalbach ; Xiaoquan Qi ; Saleha Bakht ; Abhijeet Deb Roy ; Sam T. Mugford ; Rebecca J. M. Goss ; Robert A. Field ; Anne Osbourn
J Biol Chem 2013, 288 (6) 3696-3704
Plants produce a huge array of specialized metabolites that have important functions in defense against biotic and abiotic stresses. Many of these compounds are glycosylated by family 1 glycosyltransferases (GTs). Oats (Avenaspp.) make root-derived antimicrobial triterpenes (avenacins) that provide protection against soil-borne diseases. The ability to synthesize avenacins has evolved since the divergence of oats from other cereals and grasses. The major avenacin, A-1, is acylated with N-methylanthranilic acid. Previously, we have cloned and characterized three genes for avenacin synthesis (for the triterpene synthase SAD1, a triterpene-modifying cytochrome P450 SAD2, and the serine carboxypeptidase-like acyl transferase SAD7), which form part of a biosynthetic gene cluster. Here, we identify a fourth member of this gene cluster encoding a GT belonging to clade L of family 1 (UGT74H5), and show that this enzyme is anN-methylanthranilic acid O-glucosyltransferase implicated in the synthesis of avenacin A-1. Two other closely related family 1 GTs (UGT74H6 and UGT74H7) are also expressed in oat roots. One of these (UGT74H6) is able to glucosylate both N-methylanthranilic acid and benzoic acid, whereas the function of the other (UGT74H7) remains unknown. Our investigations indicate that UGT74H5 is likely to be key for the generation of the activated acyl donor used by SAD7 in the synthesis of the major avenacin, A-1, whereas UGT74H6 may contribute to the synthesis of other forms of avenacin that are acylated with benzoic acid.
Daniel R. Tromans ; Clare E. M. Stevenson ; Rebecca J. M. Goss ; David M. Lawson
Acta Cryst 2012, F68 971-974
Pac17 is an uncharacterized protein from the pacidamycin gene cluster of the soil bacterium Streptomyces coeruleorubidus. It is implicated in the biosynthesis of the core diaminobutyric acid residue of the antibiotic, although its precise role is uncertain at present. Given that pacidamycins inhibit translocase I of Pseudomonas aeruginosa, a clinically unexploited antibiotic target, they offer new hope in the search for antibacterial agents directed against this important pathogen. Crystals of Pac17 were grown by vapour diffusion and X-ray data were collected at a synchrotron to a resolution of 1.9 Å from a single crystal. The crystal belonged to space group C2, with unit-cell parameters a = 214.12, b = 70.88, c = 142.22 Å, = 92.96°. Preliminary analysis of these data suggests that the asymmetric unit consists of one Pac17 homotetramer, with an estimated solvent content of 49.0%.
Andreas N. Tsoligkas ; James Bowen ; Michael Winn ; Rebecca J. M. Goss ; Tim W. Overton ; Mark J. H. Simmons
Colloids and Surfaces B: Biointerfaces 2012, 89 152-160
The ability of biofilms to withstand chemical and physical extremes gives them the potential to be developed as robust biocatalysts. Critical to this issue is their capacity to withstand the physical environment within a bioreactor; in order to assess this capability knowledge of their surface properties and adhesive strength is required. Novel atomic force microscopy experiments conducted under growth conditions (30 °C) were used to characterise Escherichia coli biofilms, which were generated by a recently developed spin-coating method onto a poly-l-lysine coated glass substrate. High-resolution topographical images were obtained throughout the course of biofilm development, quantifying the tip–cell interaction force during the 10 day maturation process. Strikingly, the adhesion force between the Si AFM tip and the biofilm surface increased from 0.8 nN to 40 nN within 3 days. This was most likely due to the production of extracellular polymer substance (EPS), over the maturation period, which was also observed by electron microscopy. At later stages of maturation, multiple retraction events were also identified corresponding to biofilm surface features thought to be EPS components. The spin coated biofilms were shown to have stronger surface adhesion than an equivalent conventionally grown biofilm on the same glass substrate.
Michael Winn ; Joanne M. Foulkes ; Stefano Perni ; Mark J. H. Simmons ; Tim W. Overton ; Rebecca J. M. Goss
Cat Sci Tech 2012, 2 1544-1547
The robust nature of biofilms makes them medicinally difficult to treat, however this same property renders them an attractive method for protecting and immobilising enzymes for biotransformation. Although biofilms consisting of a consortium of different microbial species have been routinely used in water purification for many decades, there are few reported examples of single species biofilms being harnessed for industrial applications. The potential of using tailored single species biofilms in order to catalyse a biotransformation of choice is attractive; we reflect upon recent advances in the use and generation of such platforms, from both biological and process engineering viewpoints.
Rebecca J. M. Goss ; Sreejith Shankar ; Antoine Abou Fayad
Nat Prod Rep 2012, 29 870-889
Natural product analogue generation is important, providing tools for chemical biology, enabling structure activity relationship determination and insight into the way in which natural products interact with their target biomolecules. The generation of analogues is also often necessary in order to improve bioavailability and to fine tune compounds’ activity. This review provides an overview of the catalogue of approaches available for accessing series of analogues. Over the last few years there have been major advances in genome sequencing and the development of tools for biosynthetic pathway engineering; it is therefore becoming increasingly easy to combine molecular biology and synthetic organic chemistry in order to enable expeditious access to series of natural products. This review outlines the various ways of combining biology and chemistry that have been applied to analogue generation, drawing upon a series of examples to illustrate each approach.
Amany E. Ragab ; Sabine Grüschow ; Daniel R. Tromans ; Rebecca J. M. Goss
JACS 2011, 133 (39) 15288-15291
The pacidamycins belong to a class of antimicrobial nucleoside antibiotics that act by inhibiting the clinically unexploited target translocase I, a key enzyme in peptidoglycan assembly. As with other nucleoside antibiotics, the pacidamycin 4′,5′-dehydronucleoside portion is an essential pharmacophore. Here we show that the biosynthesis of the pacidamycin nucleoside in Streptomyces coeruleorubidus proceeds through three steps from uridine. The transformations involve oxidation of the 5′-alcohol by Pac11, transamination of the resulting aldehyde by Pac5, and dehydration by the Cupin-domain protein Pac13.