Purine Based Combinatorial Chemistry: Solution Phase Simultaneous Addition of Functionalities. Iterative Deconvolution by Orthogonal Protection to a Single Compound with Potent Antibacterial Activity

Abstract

We have recently described a method to prepare combinatorial chemistry libraries by solution phase simultaneous addition of functionalities (SPSAF).^1–2 SPSAF has been used to create libraries based on the purine heterocycle. The nucleophilic sites (secondary nitrogens) in the planer heteroaromatic purine scaffold were built in via linkers. Thus, to continue the use of electophilic functionalities, as in previous libraries, a bifunctional nucleophilic linker was required. Piperazines readily served this purpose. Nucleophilic displacement of the chloro groups on 2,6-dichloropurine with piperazines provides reactive, constrained secondary amines for combinatorialization (Figure 1, 1). An additional piperazine was placed in the 9-position by alkylation of 2,6-dipiperazinylpurine. In this manner, the functionality that differentiates each pool (sublibrary) could be placed last in the synthetic scheme (fix last concept).¹     

Solution Phase Combinatorial Chemistry. Purine- and Pyrimidine-Based Libraries with Antibacterial Activity via Solution Phase Simultaneous Addition of Functionalities

Abstract

We have effectively used the SPSAF method with purine and pyrimidine based scaffolds to rapidly generate diverse libraries with antibacterial activity. Deconvolution via HPLC fractionation resulted in libraries with reduced complexity (2–10 compounds), a number of which retained good antibacterial activity.     

The Chemical Synthesis of E. coli tRNALys Anticodon Loop Fragment and Its Analogues

Abstract

E. coli tRNA^Lys anticodon loop fragment (Umnm⁵sUUUt⁶A) 1 and its analogues 2–6 were synthesized by the classical phosphotriester approach in solution. The preparation of suitably protected derivatives of N⁶-threonylcarbamoyladenosine 18 is also described.     

Combinatorial Generation of Nucleobase Libraries by MCR

Abstract

Multi component reactions (MCR) together with a suitable set of nucleobase building blocks provide a valuable tool for the fast generation of combinatorial libraries. These arrays of compounds are of high interest as anticancer and antiviral drugs. The preparation of small libraries and their analytics is described.     

Hydrogel composite of lanthanum and Halorubrum ejinoor sp. cell lysate as an adsorbing material

Objectives

Although halophilic archaea are rich in natural environments, their biotechnological applications are not as prevalent as those of other extremophiles, such as thermophiles and alkaliphiles. This study presents an simple method to prepare a hydrogel composite using crude cell lysate of a halophilic archaea, Halorubrum ejinoor sp. (H.e.) which was isolated from a saline lake in Inner Mongolia, China. Furthermore, formation mechanism and potential applications of the hydrogel as an adsorbing material are discussed.

Results

Halorubrum ejinoor sp. (H.e.) cell lysate was firstly prepared by adding pure water onto the H.e. cell pellet, followed by a short incubation at 60 °C. The cell lysate was injected into different metal ion (or H⁺) solutions to obtain the hydrogel composite. It was observed that H⁺, Fe³⁺, La³⁺, Cu²⁺, and Ca²⁺ induced gelation of the cell lysate, while Fe²⁺, Co²⁺, Ni²⁺, Mg²⁺, Na⁺, and K⁺ did not. DNA and extracellular polysaccharides (EPS) in the H.e. cell lysate were found to be responsible for the gelation reaction. These results suggest that DNA and EPS should be crosslinked by metal ions (or H⁺) and form a networked structure in which the metal ion (or H⁺) serves as an anchor point. Potential application of the hydrogel as an adsorbing material was explored using La³⁺-induced H.e. hydrogel composite. The hydrogel composite can adsorb the fluoride, phosphate and DNA-binding carcinogenic agents, such as acridine orange.

Conclusions

The simplicity and cost effectiveness of the preparation method might make H.e. hydrogel a promising adsorbing material. This work is expected to expand the technical applications of haloarchaea.

     

Polyphasic analysis in the description of Sulfitobacter salinus sp. nov., a marine alphaproteobacterium isolated from seawater

Purpose

A polyphasic analysis was performed on a novel bacterium, designated strain KMU-143^T, which was isolated from seawater collected in the Republic of Korea.

Methods

A novel marine bacterium KMU-143^T was analyzed and described using a polyphasic taxonomic method including 16S rRNA gene sequence analysis, DNA–DNA hybridization, and physiological, biochemical, and chemotaxonomic analyses.

Results

Strain KMU-143^T was Gram-stain-negative, strictly aerobic, oval-shaped, non-motile, and chemoorganoheterotrophic. Phylogenetic analysis based on the 16S rRNA gene sequence demonstrated that the novel marine bacterium belongs to the family Rhodobacteraceae, of the class Alphaproteobacteria, and that it possessed the highest (97.1%) sequence similarity with Sulfitobacter pontiacus ChLG 10^T and Sulfitobacter undariae W-BA2^T. DNA–DNA relatedness values between strains KMU-143^T, S. pontiacus JCM 21789^T, and S. undariae KCTC 42200^T were less than 70%. The major isoprenoid quinone of the novel isolate was ubiquinone-10 (Q-10) and the major (> 10%) cellular fatty acids were C16:0 and C18:1 ω7c. The genomic DNA G+C content of strain KMU-143^T was 56.1 mol%. The polar lipid profile of the strain KMU-143^T was found to consist of phosphatidylglycerol, diphosphatidylglycerol, phosphatidylcholine, an unidentified aminolipid, and two unidentified lipids.

Conclusion

Based on the discriminative phylogenetic position and combination of genotypic and phenotypic properties, the strain is considered to represent a new species of the genus Sulfitobacter for which the name Sulfitobacter salinus sp. nov. is proposed. The type strain of S. salinus sp. nov. is KMU-143^T (= KCCM 90322^T = NBRC 113459^T).

     

Dinucleoside Polyphosphate NAD Analogs as Potential NMN Adenylyltransferase Inhibitors. Synthesis and Biological Evaluation

Abstract

Two dinucleoside polyphosphate NAD analogs, P¹-(adenosine-5′)-P³-(nicotinamide riboside-5′)triphosphate (Np₃A, 1) and P¹-(adenosine-5′)-P⁴-(nicotinamide riboside-5′)tetraphosphate (Np₄A, 2), were synthesized and tested as inhibitors of both microbial and human recombinant NMN adenylyltransferase. Compounds 1 and 2 proved to be selective inhibitors of microbial enzymes.     

A novel amidohydrolase (DmhA) from Sphingomonas sp. that can hydrolyze the organophosphorus pesticide dimethoate to dimethoate carboxylic acid and methylamine

Objectives

To characterize a novel dimethoate amidohydrolase from Sphingomonas sp. DC-6.

Results

A gene, dmhA, encoding the dimethoate amidohydrolase responsible for transforming dimethoate to dimethoate carboxylic acid and methylamine, was cloned from Sphingomonas sp. DC-6. Sequence analysis and molecular modeling indicate that DmhA shares 31–57 % amino acid sequence identities with other functionally confirmed amidohydrolase. DmhA was expressed in Escherichia coli BL21 (DE3) and purified by Ni–NTA affinity chromatography. The purified DmhA could hydrolyze 4-acetaminophenol, dimethoate and propanil. DmhA activity was optimal at 30 °C and pH 7.5. Hg²⁺, Zn²⁺, Cu²⁺, Cd²⁺, Tween 80, Triton X-100 or SDS strongly inhibited its activity. The K _m and k _cat values of DmhA for dimethoate are 0.02 mM and 1.2 s⁻¹, respectively.

Conclusions

DmhA was confirmed to be a novel dimethoate amidohydrolase which could eliminate the toxicity of dimethoate, providing a novel gene resource for the development of pesticide-degrading enzyme preparation and mechanistic study of dimethoate hydrolysis.

     

Aflatoxin B1 and DNA Adducts. Proposed Model for Surface Noncovalent and Covalent Complexes with N(7) of Guanine. II.

Abstract

An alternate model for surface noncovalent and surface covalent binding of aflatoxin B₁ to N(7) of guanine in DNA is proposed. This model considers the out-of-plane motions of C(8) of aflatoxin B₁ in those interactions.

The covalent intercalated fit of aflatoxin B₁ into DNA arises from steric adjustments made by DNA at the covalent intercalation site as well as local strain in the bond angles about N(7) of guanine and C(8) of aflatoxin B₁. The bond angle about N(7) deviates modestly from the sp² value toward the sp³ value.

This study suggests that the surface covalent aflatoxin B₁ -DNA complex serves only a minor role in aflatoxin's precarcinogenic interaction with DNA and is a likely correctable error.     

Introduction of Thinopyrum intermedium ssp. trichophorum chromosomes to wheat by trigeneric hybridization involving Triticum,Secale and Thinopyrum genera

Main conclusion

Fluorescence in situ hybridization and molecular markers have confirmed that several chromosomes from Thinopyrum intermedium ssp. trichophorum have been added to a wheat background, which originated from a cross between a wheat– Thinopyrum partial amphiploid and triticale. The lines displayed blue grains and resistance to wheat stripe rust.

Thinopyrum intermedium has been used as a valuable resource for improving the disease resistance and yield potential of wheat. With the aim to transfer novel genetic variation from Th. intermedium species for sustainable wheat breeding, a new trigeneric hybrid was produced by crossing an octoploid wheat–Th. intermedium ssp. trichophorum partial amphiploid with hexaploid triticale. Fluorescence in situ hybridization (FISH) revealed that Thinopyrum chromosomes were transmitted preferably and the number of rye chromosomes tended to decrease gradually in the selfed derivatives of the trigeneric hybrids. Four stable wheat–Th. intermedium chromosome substitution, addition and translocation lines were selected, and a 2J^S addition line, two substitution lines of 4J^S(4B) and 4J(4B), and a small 4J.4B translocation line were identified by FISH and molecular markers. It was revealed that the gene(s) responsible for blue grains may located on the FL0.60–1.00 of long arm of Th. intermedium-derived 4J chromosome. Disease resistance screenings indicated that chromosomes 4J^S and 2J^S appear to enhance the resistance to stripe rust in the adult plant stage. The new germplasm with Th. intermedium introgression shows promise for utilization of Thinopyrum chromosome segments in future wheat improvement.

     

Plant maintenance and environmental stress. Summarising the effects of contrasting elevation,soil, and latitude on Quercus ilex respiration rates

Aims

Rice (Oryza sativa L.), wheat (Triticum aestivum L.) and common bean (Phaseolus vulgaris L.) are major staple food crops consumed worldwide. Zinc (Zn) deficiency represents a common micronutrient deficiency in human populations, especially in regions of the world where staple food crops are the main source of daily calorie intake. Foliar application of Zn fertilizer has been shown to be effective for enriching food crop grains with Zn to desirable amounts for human nutrition. For promoting adoption of this practice by growers, it is important to know whether foliar Zn fertilizers can be applied along with pesticides to wheat, rice and also common bean grown across different soil and environmental conditions.

Methods

The feasibility of foliar application of zinc sulphate (ZnSO₄.7H₂O) to wheat, rice and common bean in combination with commonly used five fungicides and nine insecticides was investigated under field conditions at the 31 sites-years of seven countries, i.e., China, India, Pakistan, Thailand, Turkey, Brazil and Zambia.

Results

Significant increases in grain yields were observed with foliar Zn/foliar Zn?+?pesticide (5.2–7.7 % of wheat and 1.6–4.2 % of rice) over yields with no Zn treatment. In wheat, as average of all experiments, higher grain Zn concentrations were recorded with foliar Zn alone (41.2 mg kg^?1) and foliar Zn?+?pesticide (38.4 mg kg^?1) as compared to no Zn treatment (28.0 mg kg^?1). Though the magnitude of grain Zn enrichment was lesser in rice than wheat, grain Zn concentrations in brown rice were significantly higher with foliar Zn (24.1 mg kg^?1) and foliar Zn?+?pesticide (23.6 mg kg^?1) than with no Zn (19.1 mg kg^?1). In case of common bean, grain Zn concentration increased from 68 to 78 mg kg^?1 with foliar Zn alone and to 77 mg kg^?1 with foliar Zn applied in combination with pesticides. Thus, grain Zn enrichment with foliar Zn, without or with pesticides, was almost similar in all the tested crops.

Conclusions

The results obtained at the 31 experimental site-years of seven countries revealed that foliar Zn fertilization can be realized in combination with commonly-applied pesticides to contribute Zn biofortification of grains in wheat, rice and common bean. This agronomic approach represents a useful practice for the farmers to alleviate Zn deficiency problem in human populations.

     

Genome-wide identification of conserved microRNA and their response to drought stress in Dongxiang wild rice (Oryza rufipogon Griff.)

Objectives

To identify drought stress-responsive conserved microRNA (miRNA) from Dongxiang wild rice (Oryza rufipogon Griff., DXWR) on a genome-wide scale, high-throughput sequencing technology was used to sequence libraries of DXWR samples, treated with and without drought stress.

Results

505 conserved miRNAs corresponding to 215 families were identified. 17 were significantly down-regulated and 16 were up-regulated under drought stress. Stem-loop qRT-PCR revealed the same expression patterns as high-throughput sequencing, suggesting the accuracy of the sequencing result was high. Potential target genes of the drought-responsive miRNA were predicted to be involved in diverse biological processes. Furthermore, 16 miRNA families were first identified to be involved in drought stress response from plants.

Conclusion

These results present a comprehensive view of the conserved miRNA and their expression patterns under drought stress for DXWR, which will provide valuable information and sequence resources for future basis studies.

     

Redefining the dose of the entomopathogenic fungus Metarhizium brunneum (Ascomycota,Hypocreales) to increase Fe bioavailability and promote plant growth in calcareous and sandy soils

Background and aims

Plants differ in their ability to use different nitrogen (N) chemical forms, these differences can be related to their ecology and drive community structure. The capacity to uptake intact organic N has been observed in plants of several ecosystems. However, soil organic N uptake by Mediterranean plants is unknown despite organic N being abundant in Mediterranean ecosystems. We compare the uptake of different N forms in two widespread coexisting Mediterranean forest trees with contrasting ecophysiological characteristics: Quercus ilex and Pinus halepensis.

Methods

To estimate root uptake rate of each N form we used equimolar solutions (1 mM N) of ¹⁵NO₃ ^?, ¹⁵NH₄ ⁺ and ¹⁵N-¹³C glycine.

Results

NH₄ ⁺ and glycine were taken up at a similar rate, but faster than NO₃ ^? in both species. Intact dual labeled glycine was found in both species, demonstrating that both species can absorb intact organic N.

Conclusions

Despite their ecological differences, both species had similar preference for N forms suggesting no fundamental niche complementarity for N uptake. The higher preference for NH₄ ⁺ and glycine over NO₃ ^? possibly reflects adaptation to the differing proportions of N forms in Mediterranean soils.

     

Tetraselmis wettsteinii (Schiller) Throndsen comb. nov. and its occurrence in golfo di Napoli

Abstract

Carteria wettsteinii Schiller (1913) has been isolated from Golfo di Napoli and investigated by means of electron microscopy which revealed micro-anatomical features characteristic of the genus Tetraselmis. The species is commonly present in Golfo di Napoli during summer with cell numbers 10²–10⁶ cells-litre^?1, occasionally producing blooms of 10⁷ cells·litre^?1.     

Interactive effects of chromate and arsenate on their uptake and speciation in Pteris ensiformis

Background and aims

Arsenate (AsV) and chromate (CrVI) inhibit each other’s uptake and translocation in As-hyperaccumulator Pteris vittata. In the present study, we extended the research to As-sensitive plant Pteris ensiformis to better understand the mechanism of their interactions.

Methods

Plants were exposed to 0, 0.75 or 7.5 mg L^?1 AsV and 0, 0.52, or 5.2 mg L^?1 CrVI for 7 d in hydroponics. Arsenic and Cr speciation were determined in nutrient solutions and plant biomass.

Results

P. ensiformis accumulated high levels of As and Cr in the rhizomes and roots with low levels in the fronds. However, P. ensiformis was much more effective in taking up Cr than As, as much more Cr was accumulated in the roots (306–6015 vs. 87–642 mg kg^?1). AsV and CrVI increased each other’s uptake in the rhizomes and roots when co-present. The AsV and CrVI taken up by P. ensiformis were reduced to arsenite (AsIII) and chromite (CrIII), possibly serving as detoxification mechanism.

Conclusions

Uptake of As and Cr induced oxidative stress as indicated by increased lipid peroxidation and electrical conductivity. Arsenic and Cr increased each other’s uptake by P. ensiformis.

     

Induction of buds in the moss Bryum pallescens Schleich. ex Schwaegr. by a metal chelate,Fe-EDTA

Abstract

Bryum pallescens remains bud-free on basal medium. Buds are induced in response to Fe-EDTA at all concentrations tested (10^?8 – 10^?4 M); 10^?4 Mbeing optimum. None of the other chelates (EDTA, Fe-EDDHA, SA) tried is effective in eliciting this response.     

Wanted: new multicomponent reactions for generating libraries of polycyclic natural products

The amalgamation of two of combinatorial chemistry's most attractive concepts--natural product libraries and multicomponent reactions (MCRs)--should provide a powerful tactic for generating libraries of bioactive compounds. Yet, despite many recent advances in this area, only a few MCRs can deliver functionalized products whose structures closely resemble that of complex polycyclic natural products. A large proportion of recently developed MCRs are based on [4+2] or [3+2] cycloadditions, and isocyanide-based processes. Because of substrate limitations, however, they are not always ideally suitable for applications in diversity-oriented synthesis of natural product-like compounds. A promising area awaiting further development is the use of transition metal-catalyzed cascade reactions.     

Nucleoside H-Phosphonates. IX. Possible Side-Reactions During Hydrogen Phosphonate Diester Formation

Abstract

Possible side-reactions during hydrogenphosphonate diester formation in the presence of various condensing reagents (e.g. arensulfonyl derivatives, chlorophosphates and acyl chlorides) have been investigated using ³¹P NMR spectroscopy.