Type III polyketide synthases (PKSs) produce an incredibly diverse group of plant specialized metabolites with medical importance despite their structural simplicity compared with the modular type I ...and II PKS systems. The type III PKSs use homodimeric proteins to construct the molecular scaffolds of plant polyketides by iterative condensations of starter and extender CoA thioesters. Ever since the structure of chalcone synthase (CHS) was disclosed in 1999, crystallographic and mutational studies of the type III PKSs have explored the intimate structural features of these enzyme reactions, revealing that seemingly minor alterations in the active site can drastically change the catalytic functions and product profiles. New structures described in this review further build on this knowledge, elucidating the detailed catalytic mechanism of enzymes that make curcuminoids, use extender substrates without the canonical CoA activator, and use noncanonical starter substrates, among others. These insights have been critical in identifying structural features that can serve as a platform for enzyme engineering via structure-guided and precursor-directed engineered biosynthesis of plant polyketides. In addition, we describe the unique properties of the recently discovered “second-generation” type III PKSs that catalyzes the one-pot formation of complex molecular scaffolds from three distinct CoA thioesters or from “CoA-free” substrates, which are also providing exciting new opportunities for synthetic biology approaches. Finally, we consider post-type III PKS tailoring enzymes, which can also serve as useful tools for combinatorial biosynthesis of further unnatural novel molecules. Recent progress in the field has led to an exciting time of understanding and manipulating these fascinating enzymes.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
2-(2-Phenylethyl)chromones (PECs) are the principal constituents contributing to the distinctive fragrance of agarwood. How PECs are biosynthesized is currently unknown. In this work, we describe a ...diarylpentanoid-producing polyketide synthase (PECPS) identified from Aquilaria sinensis. Through biotransformation experiments using fluorine-labeled substrate, transient expression of PECPS in Nicotiana benthamiana, and knockdown of PECPS expression in A. sinensis calli, we demonstrate that the C
-C
-C
scaffold of diarylpentanoid is the common precursor of PECs, and PECPS plays a crucial role in PECs biosynthesis. Crystal structure (1.98 Å) analyses and site-directed mutagenesis reveal that, due to its small active site cavity (247 Å
), PECPS employs a one-pot formation mechanism including a "diketide-CoA intermediate-released" step for the formation of the C
-C
-C
scaffold. The identification of PECPS, the pivotal enzyme of PECs biosynthesis, provides insight into not only the feasibility of overproduction of pharmaceutically important PECs using metabolic engineering approaches, but also further exploration of how agarwood is formed.
The cyanobacterial prenyltransferase AmbP3 catalyzes the reverse prenylation of the tetracyclic indole alkaloid hapalindole U at its C‐2 position. Interestingly, AmbP3 also accepts hapalindole A, a ...halogenated C‐10 epimer of hapalindole U, and catalyzes normal prenylation at its C‐2 position. The comparison of the two ternary crystal structures, AmbP3‐DMSPP/hapalindole U and AmbP3‐DMSPP/hapalindole A, at 1.65–2.00 Å resolution revealed two distinct orientations for the substrate binding that define reverse or normal prenylation. The tolerance of the enzyme for these altered orientations is attributed to the hydrophobicity of the substrate binding pocket and the plasticity of the amino acids surrounding the allyl group of the prenyl donor. This is the first study to provide the intimate structural basis for the normal and reverse prenylations catalyzed by a single enzyme, and it offers novel insight into the engineered biosynthesis of prenylated natural products.
One way or another: AmbP3 is a prenyltransferase that catalyzes both reverse and regular prenylation depending on the substrate. X‐ray crystal structure analysis of AmbP3 in complex with reverse and regular prenylation substrates elucidates the structural basis of its surprising plasticity in catalyzing two types of prenylation.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Mycolic acid-containing bacteria (MACB) are known to activate cryptic natural product biosynthesis in co-cultures with actinobacteria. We cultured Actinosynnema mirum NBRC 14064, a producer of the ...mono-cyclic polyene macrolactam mirilactam A (6), with the MACB Tsukamurella pulmonis TP-B0596. As a result, three novel compounds (mirilactams C–E, 1–3) were produced in the co-culture conditions. Compounds 1–3 were likely derived from 6 by epoxidation and subsequent spontaneous cyclization. The chemical structures and stereochemistries of 1–3 were determined by spectroscopic analyses (NMR and MS), conformational searches in the optimized potentials for liquid simulations-3 (OPLS3) force field, and calculations of electronic circular dichroism (ECD).
AmbP1 is a cyanobacterial aromatic prenyltransferase and a dedicated synthase for (R)‐3‐geranyl‐3‐isocyanovinyl indolenine (2), the biogenetic precursor for hapalindole‐type alkaloids. The ...regioselective geranylation of cis‐indolyl vinyl isonitrile (1) by the standalone AmbP1 to give 2 has been shown to require a magnesium ion (Mg2+) to suppress the formation of cis‐2‐geranylindolyl vinyl isonitrile (3). Here, we report high‐resolution crystal structures of AmbP1 in complex with 1 and geranyl S‐thiodiphosphate (GSPP) in the presence and absence of a Mg2+ effector. The comparative study of these structures revealed a unique allosteric binding site for Mg2+ that modulates the conformation of 1 in the active site of AmbP1 for its selective geranylation. This work defines the structural basis for AmbP1 catalysis in the biogenesis of hapalindole‐type alkaloids and provides the first atomic‐level insight to the allosteric regulation of prenyltransferases.
Crystal clear: High‐resolution crystal structures were obtained for ternary complexes of the prenyltransferase AmbP1 with geranyl cis‐indolyl vinyl isonitrile and geranyl S‐thiodiphosphate. Comparison of the structures obtained in the presence and absence of Mg2+ provides mechanistic insight into Mg2+‐dependent allosteric control of regiospecificity in prenyltransferases.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Ceramicines are a series of limonoids which were isolated from the bark of Malaysian
Chisocheton ceramicus
(Meliaceae) and show various biological activities. Ceramicine B, in particular, has been ...reported to show a strong lipid droplet accumulation (LDA) inhibitory activity on a mouse pre-adipocyte cell line (MC3T3-G2/PA6). With the purpose of discovering compounds with stronger activity than ceramicine B, we further investigated the constituents of
C. ceramicus
. As a result, from the bark of
C. ceramicus
four new ceramicines (ceramicines M–P,
1
–
4
) were isolated, and their structures were determined on the basis of NMR and mass spectroscopic analyses in combination with NMR chemical shift calculations. LDA inhibitory activity of
1
–
4
was evaluated. Compounds
1
–
3
showed LDA inhibitory activity, and
3
showed better selectivity than ceramicine B while showing activity at the same order of magnitude as ceramicine B. Since
3
, which possess a carbonyl group at C-7, showed better selectivity than
5
, which possess a 7α-OH group, while showing activity at the same order of magnitude as
5
, we also investigated the effect of the substituent at C-7 by synthesizing several derivatives and evaluating their LDA inhibitory activity. Accordingly, we confirmed the importance of the presence of a 7α-OH group to the LDA inhibitory activity.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
New polycyclic tetramate macrolactams, Umezawamides A (1) and B (2) were isolated from a combined-culture of Umezawaea sp. RD066910 and mycolic-acid containing bacterium Tsukamurella pulmonis ...TP-B0596. Their planar structures and partial stereochemistries were determined based on the spectroscopic analysis, MMFF conformational search, and ECD calculations. Umezawamides are the first secondary metabolites isolated from the genus Umezawaea and they exhibited cytotoxicities to P388 murine leukemia cells. Furthermore, umezawamide A (1) showed growth inhibitory activity against Candida albicans.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Cliniatines A–C (
1
–
3
), three new Amaryllidaceae alkaloids, consisting of 2,6-dimetylpyridine and lycorine-type and/or galanthamine-type were isolated from
Clivia miniata
(Lindl.) Bosse. The ...structures and absolute configurations of
1
–
3
were elucidated based on spectroscopic data and chemical correlation. Cliniatines A–C showed moderate inhibitory activity against acetylcholinesterase.
Graphic abstract
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Picrasma javanica
Blume (Simaroubaceae) is a medium-sized tree that is distributed widely in tropical Asia. In our previous study, we isolated quassinoids from
P. javanica
bark collected in Myanmar, ...and reported their antiproliferative activities. In our ongoing research for the discovery of bioactive compounds from Myanmar medicinal plants, two new quassinoids, (16
R
)-methoxyjavanicin B (
1
) and (16
S
)-methoxyjavanicin B (
2
), along with seven known compounds (
3
–
9
), were isolated during the phytochemical investigation of the CHCl
3
soluble portion of the MeOH extract of
P. javanica
wood. The structures of the new compounds were elucidated by analyses of their spectroscopic data (1D- and 2D-NMR, HRESIMS, and CD). A cytotoxicity assay revealed that compound
8
showed moderate activities against all tested cancer cell lines, the human lung (A549), breast (MCF7), and cervical (HeLa), and the normal fibroblast cell line, with IC
50
values ranging from 48.6 to 65.9 μM. Furthermore, the antibacterial assay demonstrated that
1
and
2
had the highest activities (MIC value of 1.6 μM each), followed by
5
and
3
(MIC values of 3.1 and 6.3 μM, respectively) against the Gram-positive bacterium
B. subtilis
.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
The erroneous assumption that herbal products is generally safe for consumption, is a major factor leading to the increased of herb-induced liver injury (HILI). Even though Laurus nobilis or laurel ...is a commonly used spice, the safety aspect for its consumption is under-studied. To bridge this gap of knowledge, the mutagenicity, acute toxicity, and subacute toxicity of LAURESH®, which is a standardized laurel leaf extract were evaluated. Mutagenicity study using two S. typhimurium strains, TA100 and TA98 indicated that LAURESH® does not cause base substitution and frameshift mutation, thus suggesting that LAURESH® is non-mutagenic. While acute oral toxicity on mice established the LD50 at no less than 2,000 mg/kg of body weight, and a 28-day subacute toxicity test on rat revealed the NOAEL to be 1,000 mg/kg/day. Furthermore, blood chemistry, urinalysis, necropsy, and histopathological data from subacute toxicity study on rats does not show adverse event that could be attributed to LAURESH®, thus indicating that LAURESH® is unlikely to cause HILI. Taken together, the findings from this study and previous clinical study on LAURESH®, in combination with the historic use of laurel and previous toxicity studies conducted on laurel leaves extract, strongly suggest that LAURESH® is safe for human consumption.
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