Browsing by Author "Tong, M.H."
Now showing 1 - 6 of 6
Results Per Page
Sort Options
Item Aminoacyl chain translocation catalysed by a type II thioesterase domain in an unusual non-ribosomal peptide synthetase(NATURE COMMUNICATIONS, 2022) Wang, S.; Brittain, W.D.G.; Zhang, Q.; Lu, Z.; Tong, M.H.; Wu, K.; Kyeremeh, K.; Jenner, M.; Yu, Y.; Cobb, S.L.; Deng, H.Non-Ribosomal Peptide Synthetases (NRPSs) assemble a diverse range of natural products with important applications in both medicine and agriculture. They consist of several multienzyme subunits that must interact with each other in a highly controlled manner to facilitate efficient chain transfer, thus ensuring biosynthetic fidelity. Several mechanisms for chain transfer are known for NRPSs, promoting structural diversity. Herein, we report the first biochemically characterized example of a type II thioesterase (TEII) domain capable of catalysing aminoacyl chain transfer between thiolation (T) domains on two separate NRPS subunits responsible for installation of a dehydrobutyrine moiety. Biochemical dissection of this process reveals the central role of the TEII-catalysed chain translocation event and expands the enzymatic scope of TEII domains beyond canonical (amino)acyl chain hydrolysis. The apparent co-evolution of the TEII domain with the NRPS subunits highlights a unique feature of this enzymatic cassette, which will undoubtedly find utility in biosynthetic engineering efforts.Item Discovery of a Single Monooxygenase that Catalyzes Carbamate Formation and Ring Contraction in the Biosynthesis of the Legonmycins.(Angewandte Chemie International Edition, 2015-07) Huang, S.; Tabudravu, J.; Elsayed, S.S.; Travert, J.; Peace, D.; Tong, M.H.; Kyeremeh, K.; Kelly, S.M.; Trembleau, L.; Ebel, R.; Jaspars, M.; Yu, Y.; Deng, H.Pyrrolizidine alkaloids (PAs) are a group of natural products with important biological activities. The discovery and characterization of the multifunctional FAD‐dependent enzyme LgnC is now described. The enzyme is shown to convert indolizidine intermediates into pyrrolizidines through an unusual ring expansion/contraction mechanism, and catalyze the biosynthesis of new bacterial PAs, the so‐called legonmycins. By genome‐driven analysis, heterologous expression, and gene inactivation, the legonmycins were also shown to originate from non‐ribosomal peptide synthetases (NRPSs). The biosynthetic origin of bacterial PAs has thus been disclosed for the first time.Item Identification of a fluorometabolite from Streptomyces sp. MA37: (2R3S4S)-5-fluoro-2,3,4-trihydroxypentanoic acid†(Chemical Science, 2015) Ma, L.; Bartholome, A.; Tong, M.H.; Qin, Z.; Yu, Y.; Shepherd, T.; Kyerematen, K.; Deng, H.; Hagan, D.O.(2R3S4S)-5-Fluoro-2,3,4-trihydroxypentanoic acid (5-FHPA) has been discovered as a new fluorometabolite in the soil bacterium Streptomyces sp. MA37. Exogenous addition of 5-fluoro-5-deoxy-D-ribose (5-FDR) into the cell free extract of MA37 demonstrated that 5-FDR was an intermediate to a range of unidentified fluorometabolites, distinct from fluoroacetate (FAc) and 4-fluorothreonine (4-FT). Bioinformatics analysis allowed identification of a gene cluster (fdr), encoding a pathway to the biosynthesis of 5-FHPA. Over-expression and in vitro assay of FdrC indicated that FdrC is a NAD+ dependent dehydrogenase responsible for oxidation of 5-FDR into 5-fluoro-5-deoxy-lactone, followed by hydrolysis to 5-FHPA. The identity of 5-FHPA in the fermentation broth was confirmed by synthesis of a reference compound and then co-correlation by 19F-NMR and GC-MS analysis. The occurrence of 5-FHPA proves the existence of a new fluorometabolite pathway.Item Legonoxamines A-B, two new hydroxamate siderophores from the soil bacterium, Streptomyces sp. MA37(Tetrahedron Letters, 2019-01) Maglangit, F.; Tong, M.H.; Jaspars, M.; Kyeremeh, K.; Deng, H.Two new siderophores belonging to the hydroxamate class, Legonoxamine A (1) and B (2) have been isolated from the soil bacterium, Streptomyces sp. MA37, together with one known compound, desferrioxamine B (3). Their structures were elucidated based on spectroscopic methods including 1D, 2D NMR, MS, as well as by comparison with the relevant literatures. To our knowledge, this is the first report describing a siderophore containing the N-hydroxyl phenylacetyl cadaverine (HPAC) moiety in the structure. Based on bioinformatics analysis and previous knowledge of the biosynthesis of the hydroxamate-type siderophore, the biosynthetic gene cluster (lgo) responsible for the production of 1–3 was identified in the annotated genome of the producing strain. The supplementation of phenylacetate and benzoate analogues with meta substitution into the cultures of Streptomyces sp. MA37 resulted in the production of new legonoxamine A derivatives as observed in LC-HR-ESIMS, suggesting that the legonoxamine biosynthetic pathway has a good degree of natural flexibility of accepting unnatural precursors with different functional groups.Item Targeted Dereplication of Microbial Natural Products by High-Resolution MS and Predicted LC Retention Time(Journal of Natural Products, 2017-04) Chervin, J.; Stierhof, M.; Tong, M.H.; Peace, D.; Hansen, K.O.; Urgast, D.S.; Andersen, J.H.; Yu, Y.; Ebel, R.; Kyeremeh, K.; Paget, V.; Cimpan, G.; Wyk, A.V.; Deng, H.; Jaspars, M.; Tabudravu, J.N.A new strategy for the identification of known compounds in Streptomyces extracts that can be applied in the discovery of natural products is presented. The strategy incorporates screening a database of 5555 natural products including 5098 structures from Streptomyces sp., using a high-throughput LCMS data processing algorithm that utilizes HRMS data and predicted LC retention times (tR) as filters for rapid identification of known compounds in the natural product extract. The database, named StrepDB, contains for each compound the structure, molecular formula, molecular mass, and predicted LC retention time. All identified compounds are annotated and color coded for easier visualization. It is an indirect approach to quickly assess masses (which are not annotated) that may potentially lead to the discovery of new or novel structures. In addition, a spectral database named MbcDB was generated using the ACD/Spectrus DB Platform. MbcDB contains 665 natural products, each with structure, experimental HRESIMS, MS/MS, UV, and NMR spectra. StrepDB was used to screen a mutant Streptomyces albus extract, which led to the identification and isolation of two new compounds, legonmaleimides A and B, the structures of which were elucidated with the aid of MbcDB and spectroscopic techniques. The structures were confirmed by computer-assisted structure elucidation (CASE) methods using ACD/Structure Elucidator Suite. The developed methodology suggests a pipeline approach to the dereplication of extracts and discovery of novel natural products.Item An unusual metal-bound 4-fluorothreonine transaldolase from Streptomyces sp. MA37 catalyses promiscuous transaldol reactions(Applied Microbiology and Biotechnology, 2020-03-06) Kyeremeh, K.; Wu, L.; Tong, M.H.; Raab, A.; Fang, Q.; Wang, S.; Yu, Y.; Deng, H.β-Hydroxy-α-amino acids (βH-AAs) are key components of many bioactive molecules as well as exist as specialised metabolites. Among these βH-AAs, 4-fluorothreonine (4-FT) is the only naturally occurring fluorinated AA discovered thus far. Here we report overexpression and biochemical characterisation of 4-fluorothreonine transaldolase from Streptomyces sp. MA37 (FTaseMA), a homologue of FTase previously identified in the biosynthesis of 4-FT in S. cattleya. FTaseMA displays considerable substrate plasticity to generate 4-FT as well as other β-hydroxy-α-amino acids with various functionalities at C4 position, giving the prospect of new chemo-enzymatic applications. The enzyme has a hybrid of two catalytic domains, serine hydroxymethyltransferase (S) and aldolase (A). Site-directed mutagenesis allowed the identification of the key residues of FTases, suggesting that the active site of A domain has a historical reminiscent feature in metal-dependent aldolases. Elemental analysis demonstrated that FTaseMA is indeed a Zn2+-dependent enzyme, the first example of pyridoxal phosphate (PLP) enzyme family fused with a metal-binding domain carrying out a distinct catalytic role. Finally, FTaseMA showed divergent evolutionary origin with other PLP dependent enzymes