Expression of Chromosome-integrated Hydrogen-uptake Genes in Cicer-Rhizobium

Integration of H₂-uptake (hup) gene cosmid pHU52 into the chromosomal DNA, conferred H₂- uptake activity on the Hup^- Cicer-Rhizobium strain G36–84 in the free-living state and in nodules. In five transconjugants (G36–84:: Tn5:: pHU52) derepressed for hup gene expression, the specific Hup activity ranged from 158 to 256 nmole H₂ hr^-1 mg-¹ protein which was 42 to 64% lower than the activity obtained in transconjugant with pHU52 as an episome. Integration of the cosmid significantly improved the relative efficiency of symbiotic N₂-fixation by imparting H₂-recycling capability to Hup^- Cicer-Rhizobium. Demonstration of Hup activity in the nodules of field grown chickpea plants suggests that the integrated hup genes are stably maintained in natural environment     

NAR Breakthrough Article: Next-generation sequencing reveals the biological significance of the N2,3-ethenoguanine lesion in vivo

Etheno DNA adducts are a prevalent type of DNA damage caused by vinyl chloride (VC) exposure and oxidative stress. Etheno adducts are mutagenic and may contribute to the initiation of several pathologies; thus, elucidating the pathways by which they induce cellular transformation is critical. Although N²,3-ethenoguanine (N²,3-εG) is the most abundant etheno adduct, its biological consequences have not been well characterized in cells due to its labile glycosidic bond. Here, a stabilized 2′-fluoro-2′-deoxyribose analog of N²,3-εG was used to quantify directly its genotoxicity and mutagenicity. A multiplex method involving next-generation sequencing enabled a large-scale in vivo analysis, in which both N²,3-εG and its isomer 1,N²-ethenoguanine (1,N²-εG) were evaluated in various repair and replication backgrounds. We found that N²,3-εG potently induces G to A transitions, the same mutation previously observed in VC-associated tumors. By contrast, 1,N²-εG induces various substitutions and frameshifts. We also found that N²,3-εG is the only etheno lesion that cannot be repaired by AlkB, which partially explains its persistence. Both εG lesions are strong replication blocks and DinB, a translesion polymerase, facilitates the mutagenic bypass of both lesions. Collectively, our results indicate that N²,3-εG is a biologically important lesion and may have a functional role in VC-induced or inflammation-driven carcinogenesis.     

Basis of Miscoding of the DNA Adduct N2,3-Ethenoguanine by Human Y-family DNA Polymerases

N²,3-Ethenoguanine (N²,3-ϵG) is one of the exocyclic DNA adducts produced by endogenous processes (e.g. lipid peroxidation) and exposure to bioactivated vinyl monomers such as vinyl chloride, which is a known human carcinogen. Existing studies exploring the miscoding potential of this lesion are quite indirect because of the lability of the glycosidic bond. We utilized a 2′-fluoro isostere approach to stabilize this lesion and synthesized oligonucleotides containing 2′-fluoro-N²,3-ϵ-2′-deoxyarabinoguanosine to investigate the miscoding potential of N²,3-ϵG by Y-family human DNA polymerases (pols). In primer extension assays, pol η and pol κ replicated through N²,3-ϵG, whereas pol ι and REV1 yielded only 1-base incorporation. Steady-state kinetics revealed that dCTP incorporation is preferred opposite N²,3-ϵG with relative efficiencies in the order of pol κ > REV1 > pol η ≈ pol ι, and dTTP misincorporation is the major miscoding event by all four Y-family human DNA pols. Pol ι had the highest dTTP misincorporation frequency (0.71) followed by pol η (0.63). REV1 misincorporated dTTP and dGTP with much lower frequencies. Crystal structures of pol ι with N²,3-ϵG paired to dCTP and dTTP revealed Hoogsteen-like base pairing mechanisms. Two hydrogen bonds were observed in the N²,3-ϵG:dCTP base pair, whereas only one appears to be present in the case of the N²,3-ϵG:dTTP pair. Base pairing mechanisms derived from the crystal structures explain the slightly favored dCTP insertion for pol ι in steady-state kinetic analysis. Taken together, these results provide a basis for the mutagenic potential of N²,3-ϵG.     

Amino acid substitutions of GLY98, LEU245 and GLU543 in COI1 distinctively affect jasmonate-regulated male fertility in Arabidopsis

Jasmonate (JA) regulates various plant defense and developmental processes. The F-box protein CORONATINE INSENSITIVE 1 (COI1) perceives JA signals to mediate diverse plant responses including male fertility, root growth, anthocyanin accumulation, and defense against abiotic and biotic stresses. In this study, we carried out genetic, physiological and biochemical analysis on a series of coi1 mutant alleles, and found that different amino acid mutations in COI1 distinctively affect JA-regulated male fertility in Arabidopsis. All the JA responses are disrupted by the COI1 mutations W467* in coi1-1, Q343* (coi1-6), G369E (coi1-4), G98D (coi1-5), G155E (coi1-7), D452A (coi1-9) and L490A (coi1-10), though the coi1-5 mutant (COI1_G98D) contains adequate COI1 protein (～60% of wild-type). Interestingly, the low basal level of COI1^E543K in the coi1-8 mutant (～10% of wild-type COI1 level) is sufficient for maintaining male fertility (～50% of wild-type fertility); the coi1-2 mutant with low level of COI1^L245F (～10% of wild-type) is male sterile under normal growth condition (22°C) but male fertile (～80% of wild-type fertility) at low temperature (16°C); however, both coi1-2 and coi1-8 are defective in the other JA responses (root growth, anthocyanin accumulation, and plant response to the pathogen Pst DC3000 infection).     

Experimental and DFT computational studies on 5-benzyl-4-(3,4-dimethoxyphenethyl)-2H-1,2,4-triazol-3(4H)-one

The triazole compound, 5-benzyl-4-(3,4-dimethoxyphenethyl)-2H-1,2,4-triazol-3(4H)-one, has been synthesized and characterized by ¹H-NMR, ¹³C-NMR, IR, and X-ray single-crystal determination. The compound crystallizes in the monoclinic space group P2₁ with a?=?11.8844(3) Å, b?=?17.5087(4) Å, c?=?17.3648(6) Å, β?=?99.990(2)? and Z?=?8. In addition to the molecular geometry from X-ray experiment, the molecular geometry, vibrational frequencies and gauge including atomic orbital (GIAO) ¹H- and ¹³C-NMR chemical shift values of the title compound in the ground state have been calculated using the density functional method (B3LYP) with 6-31G(d,p) basis set. The calculated results show that the optimized geometries can well reproduce the crystal structure and the theoretical vibrational frequencies and chemical shift values show good agreement with experimental ones. Besides, molecular electrostatic potential (MEP), natural bond orbital (NBO), and frontier molecular orbitals (FMO) analysis of the title compound were performed by the B3LYP/6-31G(d,p) method.     

Thermodynamic Characterization of ppGpp Binding to EF-G or IF2 and of Initiator tRNA Binding to Free IF2 in the Presence of GDP, GTP, or ppGpp

In addition to their natural substrates GDP and GTP, the bacterial translational GTPases initiation factor (IF) 2 and elongation factor G (EF-G) interact with the alarmone molecule guanosine tetraphosphate (ppGpp), which leads to GTPase inhibition. We have used isothermal titration calorimetry to determine the affinities of ppGpp for IF2 and EF-G at a temperature interval of 5-25 °C. We find that ppGpp has a higher affinity for IF2 than for EF-G (1.7-2.8 μM K_dversus 9.1-13.9 μM K_d at 10-25 °C), suggesting that during stringent response in vivo, IF2 is more responsive to ppGpp than to EF-G. We investigated the effects of ppGpp, GDP, and GTP on IF2 interactions with fMet-tRNA^fMet demonstrating that IF2 binds to initiator tRNA with submicromolar K_d and that affinity is altered by the G nucleotides only slightly. This—in conjunction with earlier reports on IF2 interactions with fMet-tRNA^fMet in the context of the 30S initiation complex, where ppGpp was suggested to strongly inhibit fMet-tRNA^fMet binding and GTP was suggested to strongly promote fMet-tRNA^fMet binding—sheds new light on the mechanisms of the G-nucleotide-regulated fMet-tRNA^fMet selection.     

Experimental and theoretical investigation of the molecular and electronic structure of 5-(4-aminophenyl)-4-(3-methyl-3-phenylcyclobutyl)thiazol-2-amine

The title molecule, 5-(4-aminophenyl)-4-(3-methyl-3-phenylcyclobutyl)thiazol-2-amine (C₂₀H₂₁N₃S), was prepared and characterized by ¹H-NMR, ¹³C-NMR, IR and single-crystal X-ray diffraction. The compound crystallizes in the monoclinic space group P2₁/c with a?=?9.4350(5) Å, b?=?11.2796(6) Å, c?=?18.4170(8) Å and β?=?113.378(3)°. In addition to the molecular geometry from X-ray experiment, the molecular geometry, vibrational frequencies, gauge including atomic orbital (GIAO) ¹H- and ¹³C-NMR chemical shift values and atomic charges distribution of the title compound in the ground state have been calculated using the Hartree–Fock (HF) and density functional method (DFT) (B3LYP) with 6-31G(d) basis set. To determine conformational flexibility, molecular energy profile of the title compound was obtained by semi-empirical (AM1) calculations with respect to two selected degrees of torsional freedom, which were varied from ?180° to +180° in steps of 10°. Besides, frontier molecular orbitals (FMO) analysis was performed by the B3LYP/6-31G(d) method.     

An unexpectedly lichenase-stable hexasaccharide from cereal,horsetail and lichen mixed-linkage β-glucans (MLGs): Implications for MLG subunit distribution

Mixed-linkage (1→3),(1→4)-β-d-glucan (MLG) is a biologically and technologically important hemicellulose, known to occur in three widely separated lineages: the Poales (including grasses and cereals), Equisetum (fern-allies), and some lichens e.g. Iceland moss (Cetraria islandica). Lichenase (E.C. 3.2.1.73) is widely assumed to hydrolyse all (1→4) bonds that immediately follow (1→3) bonds in MLG, generating predominantly the tetrasaccharide β-d-Glcp-(1→4)-β-d-Glcp-(1→4)-β-d-Glcp-(1→3)-d-Glc (G4G4G3G; MLG4), the corresponding trisaccharide (G4G3G; MLG3), and sometimes also laminaribiose (G3G; MLG2). The ratio of the oligosaccharides produced characterises each polysaccharide. We report here that digestion of MLG from barley (Hordeum vulgare), Equisetum arvense and C. islandica by Bacillus subtilis lichenase also yields the unexpectedly stable hexasaccharide, β-d-Glcp-(1→3)-β-d-Glcp-(1→4)-β-d-Glcp-(1→4)-β-d-Glcp-(1→4)-β-d-Glcp-(1→3)-d-Glc (G3G4G4G4G3G, i.e. MLG2–MLG4), identified by thin-layer chromatography, gel-permeation chromatography, HPLC (HPAEC), β-glucosidase digestion, ¹H/¹³C-NMR spectroscopy and mass spectrometry. On HPLC, G3G4G4G4G3G is the major constituent of a peak previously ascribed solely to the nonasaccharide G4G4G4G4G4G4G4G3G. Because it was widely presumed that lichenase would cleave G3G4G4G4G3G to MLG2 + MLG4, our data both redefine the substrate specificity of Bacillus lichenase and show previous attempts to characterise MLGs by HPLC of lichenase-digests to be flawed. MLG2 subunits are particularly underestimated; often reported as negligible, they are here shown to be an appreciable constituent of MLGs from all three lineages. We also show that there is no appreciable yield of water-soluble lichenase products with DP > 9; potential identities of products previously labelled DP > 9 are suggested. Finally, this discovery also provides a opportunity to investigate the spatial distribution of subunits along the MLG chain. We show that MLG2 subunits in barley and Cetraria MLG are not randomly distributed, but predominantly found at the non-reducing end of MLG4 subunits.     

Effects of age and sex on the induction by triiodothyronine of cortisone delta4-5alpha-reductase and glucose 6-phosphate dehydrogenase of rat liver and adrenal

The effects of age and sex on the induction by 3,5,3′-triiodothyronine (T-3) of cortisone Δ⁴-5α-reductase and glucose 6-phosphate dehydrogenase (G 6-P D) in liver and the latter in adrenal have been investigated. Levels of cortisone Δ⁴-(5α, 5β)-reductase and G 6-P D were measured in homogenates of tissue from normal and T-3 injected male and female rats, $\text{1 1}\text{4}$ to 21 months of age. Increases in the levels of the reductase seen under T-3 stimulation were ascribed to induction of the 5α-reductase alone. T-3 caused induction of cortisone Δ⁴-5α-reductase only in the livers of male rats $\text{1}\text{3}\text{4}$ months of age. There was induction of total G 6-P D at most ages except in the livers of old male and young female rats and adrenals of young and old male rats. At all ages in normal animals of both sexes the maximum activity of glucose 6-phosphate dehydrogenase was much greater than that of cortisone Δ⁴-(5α, 5β)-reductase. It is concluded that the amount of G 6-P D in normal liver may be sufficient to handle an increase in cortisone reduction, and factors other than cortisone Δ⁴-reductase or G 6-P D levels alone must regulate increased reduction of the steroid.     

Stimulus Bias Provides Evidence for Conformational Constraints in the Structure of a G Protein-coupled Receptor

A key characteristic of G protein-coupled receptors (GPCRs) is that they activate a plethora of signaling pathways. It is now clear that a GPCR coupling to these pathways can be regulated selectively by ligands that differentially drive signaling down one pathway in preference to another. This concept, termed stimulus bias, is revolutionizing receptor biology and drug discovery by providing a means of selectively targeting receptor signaling pathways that have therapeutic impact. Herein, we utilized a novel quantitative method that determines stimulus bias of synthetic GPCR ligands in a manner that nullifies the impact of both the cellular background and the “natural bias” of the endogenous ligand. By applying this method to the M₂ muscarinic acetylcholine receptor, a prototypical GPCR, we found that mutation of key residues (Tyr-80^2.61 and Trp-99^3.28) in an allosteric binding pocket introduces stimulus bias in response to the atypical ligands AC-42 (4-n-butyl-1-(4-(2-methylphenyl)-4-oxo-1-butyl)piperidine HCl) and 77-LH-28-1 (1-(3-(4-butyl-1-piperidinyl)propyl)- 3,3-dihydro-2(1H)-quinolinone). By comparing stimulus bias factors among receptor internalization, G protein activation, extracellular-regulated protein kinase 1/2 (ERK1/2) signaling, and receptor phosphorylation, we provide evidence that Tyr-80^2.61 and Trp-99^3.28 act either as molecular switches or as gatekeeper residues that introduce constraints limiting the active conformation of the M₂ muscarinic acetylcholine receptor and thereby regulate stimulus bias. Furthermore, we provide evidence that downstream signaling pathways previously considered to be related to each other (i.e. receptor phosphorylation, internalization, and activation of ERK1/2) can act independently.     

Novel metabolites from the roots of Cadaba natalensis

From an extract of the roots of Cadaba natalensis Sond. the novel macrocyclic dibenzo-diazacyclododecanedione 1 was isolated together with (S)-2-ethyl-2-methyloxazolidin-5-one (2a). In addition, the five known compounds were obtained. Of these, (R)-5-ethyl-5-methyloxazolidin-2-one (3) is reported as a natural product for the first time. The structures of the isolated compounds were elucidated by analysis of the spectral data, 1D NMR (¹H, ¹³C, and DEPT), 2D NMR (COSY, HMQC, HMBC, and NOESY), and HRESIMS, as well as by the comparison with previously reported data.     

PAI-1 4G/5G Polymorphism Contributes to Cancer Susceptibility: Evidence from Meta-Analysis

Background

The plasminogen activator inhibitor-1 (PAI-1) is expressed in many cancer cell types and allows the modulation of cancer growth, invasion and angiogenesis. To date, studies investigated the association between a functional polymorphism in PAI-1 (4G/5G) and risk of cancer have shown inclusive results.

Methods

A meta-analysis based on 25 case-control studies was performed to address this issue. Odds ratios (OR) with corresponding 95% confidence intervals (CIs) were used to assess the association. The statistical heterogeneity across studies was examined with I² test.

Results

Overall, a significant increased risk of cancer was associated with the PAI-1 4G/4G polymorphism for the allele contrast (4G vs. 5G: OR = 1.10, CI = 1.03–1.18, I² = 49.5%), the additive genetic model (4G/4G vs. 5G/5G: OR = 1.21, CI = 1.06–1.39, I² = 51.9%), the recessive genetic model (4G/4G vs. 4G/5G+5G/5G: OR = 1.11, CI = 1.04–1.18, I² = 20.8%). In the subgroup analysis by ethnicity, the results indicated that individuals with 4G/4G genotype had a significantly higher cancer risk among Caucasians (4G/4G vs. 5G/5G: OR = 1.31, 95%CI = 1.09–1.59, I² = 59.6%; 4G/4G vs. 4G/5G: OR = 1.12, 95%CI = 1.04–1.21, I² = 3.6%; recessive model: OR = 1.12, 95%CI = 1.05–1.21, I² = 25.3%).

Conclusions

The results of the present meta-analysis support an association between the PAI-1 4G/5G polymorphism and increasing cancer risk, especially among Caucasians, and those with 4G allele have a high risk to develop colorectal cancer and endometrial cancer.     

APOBEC3G Restricts HIV-1 to a Greater Extent than APOBEC3F and APOBEC3DE in Human Primary CD4+ T Cells and Macrophages

APOBEC3 proteins inhibit HIV-1 replication in experimental systems and induce hypermutation in infected patients; however, the relative contributions of several APOBEC3 proteins to restriction of HIV-1 replication in the absence of the viral Vif protein in human primary CD4⁺ T cells and macrophages are unknown. We observed significant inhibition of HIV-1Δvif produced in 293T cells in the presence of APOBEC3DE (A3DE), APOBEC3F (A3F), APOBEC3G (A3G), and APOBEC3H haplotype II (A3H HapII) but not APOBEC3B (A3B), APOBEC3C (A3C), or APOBEC3H haplotype I (A3H HapI). Our previous studies showed that Vif amino acids Y⁴⁰RHHY⁴⁴ are important for inducing proteasomal degradation of A3G, whereas amino acids ¹⁴DRMR¹⁷ are important for degradation of A3F and A3DE. Here, we introduced substitution mutations of ⁴⁰YRHHY⁴⁴ and ¹⁴DRMR¹⁷ in replication-competent HIV-1 to generate vif mutants NL4-3 YRHHY>A5 and NL4-3 DRMR>A4 to compare the antiviral activity of A3G to the combined antiviral activity of A3F and A3DE in activated CD4⁺ T cells and macrophages. During the first 15 days (round 1), in which multiple cycles of viral replication occurred, both the NL4-3 YRHHY>A5 and NL4-3 DRMR>A4 mutants replicated in activated CD4⁺ T cells and macrophages, and only the NL4-3 YRHHY>A5 mutant showed a 2- to 4-day delay in replication compared to the wild type. During the subsequent 27 days (round 2) of cultures initiated with peak virus obtained from round 1, the NL4-3 YRHHY>A5 mutant exhibited a longer, 8- to 10-day delay and the NL4-3 DRMR>A4 mutant exhibited a 2- to 6-day delay in replication compared to the wild type. The NL4-3 YRHHY>A5 and NL4-3 DRMR>A4 mutant proviruses displayed G-to-A hypermutations primarily in GG and GA dinucleotides as expected of A3G- and A3F- or A3DE-mediated deamination, respectively. We conclude that A3G exerts a greater restriction effect on HIV-1 than A3F and A3DE.     

Novel Genetic Locus Implicated for HIV-1 Acquisition with Putative Regulatory Links to HIV Replication and Infectivity: A Genome-Wide Association Study

Fifty percent of variability in HIV-1 susceptibility is attributable to host genetics. Thus identifying genetic associations is essential to understanding pathogenesis of HIV-1 and important for targeting drug development. To date, however, CCR5 remains the only gene conclusively associated with HIV acquisition. To identify novel host genetic determinants of HIV-1 acquisition, we conducted a genome-wide association study among a high-risk sample of 3,136 injection drug users (IDUs) from the Urban Health Study (UHS). In addition to being IDUs, HIV- controls were frequency-matched to cases on environmental exposures to enhance detection of genetic effects. We tested independent replication in the Women’s Interagency HIV Study (N=2,533). We also examined publicly available gene expression data to link SNPs associated with HIV acquisition to known mechanisms affecting HIV replication/infectivity. Analysis of the UHS nominated eight genetic regions for replication testing. SNP rs4878712 in FRMPD1 met multiple testing correction for independent replication (P=1.38x10^-4), although the UHS-WIHS meta-analysis p-value did not reach genome-wide significance (P=4.47x10^-7 vs. P<5.0x10^-8) Gene expression analyses provided promising biological support for the protective G allele at rs4878712 lowering risk of HIV: (1) the G allele was associated with reduced expression of FBXO10 (r=-0.49, P=6.9x10^-5); (2) FBXO10 is a component of the Skp1-Cul1-F-box protein E3 ubiquitin ligase complex that targets Bcl-2 protein for degradation; (3) lower FBXO10 expression was associated with higher BCL2 expression (r=-0.49, P=8x10^-5); (4) higher basal levels of Bcl-2 are known to reduce HIV replication and infectivity in human and animal in vitro studies. These results suggest new potential biological pathways by which host genetics affect susceptibility to HIV upon exposure for follow-up in subsequent studies.     

Locating the uracil-5-yl radical formed upon photoirradiation of 5-bromouracil-substituted DNA

In a previous study, we found that 2-deoxyribonolactone is effectively generated in the specific 5-bromouracil (^BrU)-substituted sequence 5′-(G/C)[A]_{n = 1,2}^BrU^BrU-3′ and proposed that a formed uracil-5-yl radical mainly abstracts the C1′ hydrogen from the 5′-side of ^BrU^BrU under 302-nm irradiation condition. In the present work, we performed photoirradiation of ^BrU-substituted DNA in the presence of a hydrogen donor, tetrahydrofuran, to quench the uracil-5-yl radical to uracil and then subjected the sample to uracil DNA glycosylase digestion. Slab gel sequence analysis indicated that uracil residues were formed at the hot-spot sequence of 5′-(G/C)[A]_{n = 1,2}^BrU^BrU-3′ in 302-nm irradiation of ^BrU-substituted DNA. Furthermore, we found that the uracil residue was also formed at the reverse sequence 5′-^BrU^BrU[A]_{n = 1,2}(G/C)-3′, which suggests that both 5′-(G/C)[A]_{n = 1,2}^BrU^BrU-3′ and 5′-^BrU^BrU[A]_{n = 1,2}(G/C)-3′ are hot-spot sequences for the formation of the uracil-5-yl radical.     

An experimental and theoretical approach to the molecular structure of 2-{4-[3-(2,5-dimethylphenyl)-3-methylcyclobutyl]thiazol-2-yl}isoindoline-1,3-dione

The title compound, 2-{4-[3-(2,5-dimethylphenyl)-3-methylcyclobutyl]thiazol-2-yl}isoindoline-1,3-dione (C₂₄H₂₂N₂O₂S), was synthesized and characterized by IR-NMR spectroscopy and single-crystal X-ray diffraction. The compound crystallizes in the monoclinic space group P2₁/c with a?=?19.7799(13) Å, b?=?6.7473(4) Å, c?=?15.7259(9) Å and β?=?103.416(5)°. In addition, the molecular geometry, vibrational frequencies and gauge including atomic orbital (GIAO) ¹H and ¹³C chemical shift values of the title compound in the ground state have been calculated by using the Hartree-Fock (HF) and density functional method (DFT/B3LYP) with 6–31G(d), 6–31 + G(d,p) and LANL2DZ basis sets, and compared with the experimental data. To determine conformational flexibility, molecular energy profile of the title compound was obtained by semi-empirical (AM1) calculations with respect to two selected degrees of torsional freedom, which were varied from ?180° to +180° in steps of 5°. Besides, molecular electrostatic potential, frontier molecular orbitals (FMO) analysis and thermodynamic properties of the title compound were investigated by theoretical calculations.     

Plant regeneration from cotyledon protoplasts of glycine canescens and G. clandestina

Protoplasts isolated enzymatically from cotyledons of wild Glycine species were cultured in agarose-solidified droplets of Kao protoplast culture medium. Incubation of droplets in liquid Kao medium mixed with a B5-based formulation resulted in protoplast-derived colonies. Subculture of tissues with liquid SC2 medium containing B5 salts and vitamins, 3% sucrose, 1.1 mg 1⁻¹ BAP and 0.005 mg 1⁻¹ IBA, followed by transfer to agar solidified SC2 medium, induced shoots in two accessions, G. clandestina G1231 and G. canescens G1171. Shoots elongated on agar B5-based medium with 0.2 mg 1⁻¹ BAP and 0.005 mg 1⁻¹ IBA(SC6), and rooted on hormone-free, half-strength semi-solid B5 medium.     

Synthesis, crystal structure, NMR and ab initio molecular-orbital studies of some magnesium(II) macrocyclic Schiff-base complexes, with two 2-aminoethyl pendant arms

Three new Mg(II) bis(pendant arm) macrocyclic Schiff-base complexes, [MgLⁿ]²⁺(n=5, 6, 7), have been prepared via cyclocondensation of 2,6-diacetylpyridine with branched hexaamines and characterised spectroscopically. In addition, for [MgL⁵](ClO₄)₂ the crystal structure is reported. This is the first X-ray structural determination of an Mg(II) complex coordinated by seven nitrogen atoms. The ligands, L, are 15-, 16- and 17-membered pentaaza macrocycles having two 2-aminoethyl pendant arms [L⁵; 2,13-dimethyl-6,9-bis(aminoethyl)-3,6,9,12,18-pentaazabicyclo[12.3.1]octadeca-1(18), 2, 12, 14, 16-pentaene, L⁶; 2,14-dimethyl-6,10-bis(aminoethyl)-3,6,10,13,19-pentaazabicyclo[13.3.1]nonadeca-1(19), 2, 13, 15, 17-pentaene and L⁷; 2,15-dimethyl-6,11-bis(aminoethyl)-3,6,11,14,20-pentaazabicyclo[14.3.1]eicosa-1(20),2,14,16,18-pentaene]. The crystal structure of [MgL⁵](ClO₄)₂, was determined by X-ray diffraction and showed that the complex cation that had formed consisted of a pentagonal bipyramidally coordinated Mg(II) ion. All complexes were characterised by IR, ¹H NMR,¹³C NMR, COSY(H,H) and HETCOR(H,C) spectroscopy, and the data indicate that the structure is approximately pentagonal bipyramidal in each case. This structural assignment is also supported by ab initio HF-MO calculations made using the standard 3-21G* basis set.