Antitumor pyrrolobenzodiazepines (PBDs), lincosamide antibiotics, quorum-sensing molecule hormaomycin, and antimicrobial griselimycin are structurally and functionally diverse groups of actinobacterial metabolites. The common feature of these compounds is the incorporation of l-tyrosine- or l-leucine-derived 4-alkyl-l-proline derivatives (APDs) in their structures. Here, we report that the last reaction in the biosynthetic pathway of APDs, catalyzed by F420H2-dependent Apd6 reductases, contributes to the structural diversity of APD precursors. Specifically, the heterologous overproduction of six Apd6 enzymes demonstrated that Apd6 from the biosynthesis of PBDs and hormaomycin can reduce only an endocyclic imine double bond, whereas Apd6 LmbY and partially GriH from the biosyntheses of lincomycin and griselimycin, respectively, also reduce the more inert exocyclic double bond of the same 4-substituted Δ1-pyrroline-2-carboxylic acid substrate, making LmbY and GriH unusual, if not unique, among reductases. Furthermore, the differences in the reaction specificity of the Apd6 reductases determine the formation of the fully saturated APD moiety of lincomycin versus the unsaturated APD moiety of PBDs, providing molecules with optimal shapes to bind their distinct biological targets. Moreover, the Apd6 reductases establish the first F420H2-dependent enzymes from the luciferase-like hydride transferase protein superfamily in the biosynthesis of bioactive molecules. Finally, our bioinformatics analysis demonstrates that Apd6 and their homologues, widely distributed within several bacterial phyla, play a role in the formation of novel yet unknown natural products with incorporated l-proline-like precursors and likely in the microbial central metabolism.
- MeSH
- benzodiazepiny chemie metabolismus farmakologie MeSH
- cyklické peptidy biosyntéza chemie farmakologie MeSH
- depsipeptidy biosyntéza chemie farmakologie MeSH
- katalýza MeSH
- linkomycin biosyntéza chemie farmakologie MeSH
- molekulární modely MeSH
- oxidoreduktasy chemie metabolismus MeSH
- prolin analogy a deriváty metabolismus MeSH
- pyrroly chemie metabolismus farmakologie MeSH
- riboflavin analogy a deriváty chemie metabolismus MeSH
- substrátová specifita MeSH
- tyrosin analogy a deriváty metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- benzodiazepiny MeSH
- coenzyme F420 MeSH Prohlížeč
- cyklické peptidy MeSH
- depsipeptidy MeSH
- F420H2 dehydrogenase MeSH Prohlížeč
- griselimycin MeSH Prohlížeč
- hormaomycin MeSH Prohlížeč
- linkomycin MeSH
- oxidoreduktasy MeSH
- prolin MeSH
- pyrrolo(2,1-c)(1,4)benzodiazepine MeSH Prohlížeč
- pyrroly MeSH
- riboflavin MeSH
- tyrosin MeSH
Natural pyrrolobenzodiazepines (PBDs) form a large and structurally diverse group of antitumour microbial metabolites produced through complex pathways, which are encoded within biosynthetic gene clusters. We sequenced the gene cluster of limazepines and proposed their biosynthetic pathway based on comparison with five available gene clusters for the biosynthesis of other PBDs. Furthermore, we tested two recombinant proteins from limazepine biosynthesis, Lim5 and Lim6, with the expected substrates in vitro. The reactions monitored by LC-MS revealed that limazepine biosynthesis involves a new way of 3-hydroxyanthranilic acid formation, which we refer to as the chorismate/DHHA pathway and which represents an alternative to the kynurenine pathway employed for the formation of the same precursor in the biosynthesis of other PBDs. The chorismate/DHHA pathway is presumably also involved in the biosynthesis of PBD tilivalline, several natural products unrelated to PBDs, and its part is shared also with phenazine biosynthesis. The similarities between limazepine and phenazine biosynthesis indicate tight evolutionary links between these groups of compounds.
- MeSH
- benzodiazepiny chemie metabolismus MeSH
- chromatografie kapalinová MeSH
- hmotnostní spektrometrie MeSH
- kyselina 3-hydroxyanthranilová metabolismus MeSH
- metabolické sítě a dráhy MeSH
- molekulární evoluce MeSH
- sekvenční analýza proteinů MeSH
- Streptomyces genetika metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- benzodiazepiny MeSH
- kyselina 3-hydroxyanthranilová MeSH
Covering: up to 2017This review covers the biosynthetic and evolutionary aspects of lincosamide antibiotics, antitumour pyrrolobenzodiazepines (PBDs) and the quorum-sensing molecule hormaomycin. These structurally and functionally diverse groups of complex natural products all incorporate rarely occurring 4-alkyl-l-proline derivatives (APDs) biosynthesized from l-tyrosine through an unusual specialized pathway catalysed by a common set of six proteins named Apd1-Apd6. We give an overview of APD formation, which involves unusual enzyme activities, and its incorporation, which is based either on nonribosomal peptide synthetase (PBDs, hormaomycin) or a unique hybrid ergothioneine-dependent condensation system followed by mycothiol-dependent sulphur atom incorporation (lincosamides). Furthermore, within the public databases, we identified 36 novel unannotated biosynthetic gene clusters that putatively encode the biosynthesis of APD compounds. Their products presumably include novel PBDs, but also novel classes of APD compounds, indicating an unprecedented potential for the diversity enhancement of these functionally versatile complex metabolites. In addition, phylogenetic analysis of known and novel gene clusters for the biosynthesis of APD compounds allowed us to infer novel evolutionary hypotheses: Apd3 methyltransferase originates from a duplication event in a hormaomycin biosynthetic gene cluster ancestor, while putative Apd5 isomerase is evolutionarily linked to PhzF protein from the biosynthesis of phenazines. Lastly, we summarize the achievements in preparing hybrid APD compounds by directing their biosynthesis, and we propose that the number of nature-like APD compounds could by multiplied by replacing l-proline residues in various groups of complex metabolites with APD, i.e. by imitating the natural process that occurs with lincosamides and PBDs, in which the replacement of l-proline for APD has proved to be an evolutionary successful concept.
- MeSH
- antibakteriální látky chemie farmakologie MeSH
- benzodiazepiny chemie farmakologie MeSH
- biologické přípravky chemie metabolismus farmakologie MeSH
- cystein metabolismus MeSH
- depsipeptidy chemie metabolismus farmakologie MeSH
- ergothionein metabolismus MeSH
- glykopeptidy metabolismus MeSH
- inositol metabolismus MeSH
- lidé MeSH
- linkomycin chemie farmakologie MeSH
- linkosamidy biosyntéza farmakologie MeSH
- molekulární evoluce * MeSH
- molekulární struktura MeSH
- protinádorové látky chemie metabolismus farmakologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
- Názvy látek
- antibakteriální látky MeSH
- benzodiazepiny MeSH
- biologické přípravky MeSH
- cystein MeSH
- depsipeptidy MeSH
- ergothionein MeSH
- glykopeptidy MeSH
- hormaomycin MeSH Prohlížeč
- inositol MeSH
- linkomycin MeSH
- linkosamidy MeSH
- mycothiol MeSH Prohlížeč
- protinádorové látky MeSH
Herein, we report a stereoselective formation of tetrahydro-6H-benzo[e][1,4]oxazino[4,3-a][1,4]diazepine-6,12(11H)-diones. Their preparation consisted in solid-phase synthesis of linear intermediates starting from polymer-supported Ser(tBu)-OH. Using various 2-nitrobenzoic acids and bromoketones, the key intermediates were obtained in five steps and subjected to trifluoroacetic acid-mediated cleavage from the resin, followed by stereoselective reduction with triethylsilane. Subsequent catalytic hydrogenation of the nitro group and cyclization yielded the target compounds with full retention of the C12a stereocenter configuration.
- Klíčová slova
- benzodiazepine, morpholine, oxazine, solid-phase synthesis, stereoselective synthesis, triethylsilane,
- MeSH
- benzodiazepiny chemická syntéza chemie MeSH
- cyklizace MeSH
- katalýza MeSH
- molekulární struktura MeSH
- oxaziny chemická syntéza chemie MeSH
- stereoizomerie MeSH
- techniky kombinatorické chemie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- benzodiazepiny MeSH
- oxaziny MeSH
Anticancer pyrrolobenzodiazepines (PBDs) are one of several groups of natural products that contain unusual 4-alkyl-l-proline derivatives (APDs) in their structure. APD moieties of PBDs are characterized by high structural diversity achieved through unknown biosynthetic machinery. Based on LC-MS analysis of culture broths, feeding experiments, and protein assays, we show that APDs are not incorporated into PBDs in their final form as was previously hypothesized. Instead, a uniform building block, 4-propylidene-l-proline or 4-ethylidene-l-proline, enters the condensation reaction. The subsequent postcondensation steps are initiated by the introduction of an additional double bond catalyzed by a FAD-dependent oxidoreductase, which we demonstrated with Orf7 from anthramycin biosynthesis. The resulting double bond arrangement presumably represents a prerequisite for further modifications of the APD moieties. Our study gives general insight into the diversification of APD moieties of natural PBDs and provides proof-of-principle for precursor directed and combinatorial biosynthesis of new PBD-based antitumor compounds.
- MeSH
- benzodiazepinony chemie MeSH
- benzodiazepiny chemie klasifikace MeSH
- biologické přípravky chemie metabolismus MeSH
- farmaceutická chemie MeSH
- molekulární struktura MeSH
- protinádorové látky chemie metabolismus MeSH
- pyrroly chemie klasifikace MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- benzodiazepinony MeSH
- benzodiazepiny MeSH
- biologické přípravky MeSH
- protinádorové látky MeSH
- pyrrolo(2,1-c)(1,4)benzodiazepine MeSH Prohlížeč
- pyrroly MeSH
- tomaymycin MeSH Prohlížeč
Solid-phase synthesis of 1,2,3,4-tetrahydro-benzo[e][1,4]diazepin-5-ones with use of polystyrene resin is described. The starting material was polymer supported 1,2-diaminoethane and as a key synthon, 4-chloro-2-fluoro-5-nitrobenzoic acid was used. The synthetic approach allows the preparation of derivatives with variable substitution at positions 4 and 8. Additionally, a skeletal diversity was increased when the nitro group was reduced and some benzene fused heterocycles were prepared. An expansion of a diazepinone to a benzodiazocinone scaffold was also successful although some limitations in a diversity of target derivatives were observed.
HIV protease is a primary target for the design of virostatics. Screening of libraries of non-peptide low molecular weight compounds led to the identification of several new compounds that inhibit HIV PR in the low micromolar range. X-ray structure of the complex of one of them, a dibenzo[b,e][1,4]diazepinone derivative, showed that two molecules of the inhibitor bind to the PR active site. Covalent linkage of two molecules of such a compound by a two-carbon linker led to a decrease of the inhibition constant of the resulting compound by 3 orders of magnitude. Molecular modeling shows that these dimeric inhibitors form two crucial hydrogen bonds to the catalytic aspartates that are responsible for their improved activity compared to the monomeric parental building blocks. Dibenzo[b,e][1,4]diazepinone analogues might represent a potential new class of HIV PIs.
- MeSH
- benzodiazepiny chemie MeSH
- HIV infekce farmakoterapie enzymologie virologie MeSH
- HIV-1 účinky léků MeSH
- HIV-proteasa chemie metabolismus MeSH
- inhibitory HIV-proteasy chemická syntéza farmakologie MeSH
- katalytická doména MeSH
- katalýza MeSH
- konformace proteinů MeSH
- krystalografie rentgenová MeSH
- lidé MeSH
- molekulární modely MeSH
- molekulární struktura MeSH
- peptidové fragmenty farmakologie MeSH
- racionální návrh léčiv * MeSH
- vodíková vazba MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- benzodiazepiny MeSH
- HIV-proteasa MeSH
- inhibitory HIV-proteasy MeSH
- peptidové fragmenty MeSH
The problems of the concentration dependence of retention indices and the applicability of extrapolated values in the evaluation of lipophilicity were studied. The reversed-phase high-performance liquid chromatography of arylalkanoic acids were carried out with experimental data for substituted estra-1,3,5 (10)-trienes, benzodiazepines, dermorphine derivatives and dansylamides selected from the literature for this purpose. Fair linear relationships between slopes of concentration dependences and extrapolated and non-extrapolated values of RM and log k' were found. Equivalence of these indices in the evaluation of lipophilicity can be inferred. Statistically significant dependences of log P (sigma pi) values on concentration slopes make it possible to use them as new parameters of lipophilicity. The goodness of fit of these relationships increases when the values of ET(30), as a measure of the solvatochromic solvent polarity of mobile phases, are used instead of the change in modifier concentration.
- MeSH
- acetáty chemie MeSH
- benzodiazepiny chemie MeSH
- chemické jevy MeSH
- chromatografie na tenké vrstvě MeSH
- chromatografie * MeSH
- fyzikální chemie MeSH
- lipidy * MeSH
- molekulární sekvence - údaje MeSH
- oligopeptidy chemie MeSH
- opioidní peptidy MeSH
- sekvence aminokyselin MeSH
- sloučeniny dansylu chemie MeSH
- vysokoúčinná kapalinová chromatografie MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- acetáty MeSH
- benzodiazepiny MeSH
- dermorphin MeSH Prohlížeč
- lipidy * MeSH
- oligopeptidy MeSH
- opioidní peptidy MeSH
- sloučeniny dansylu MeSH
