Localization of repetitive and unique DNA sequences neighbouring the rabbit beta-globin gene

The structure of oxymyoglobin has been refined at 1·6 Å resolution, using diffractometer data collected at ?12 °C. The crystallographic R factor is 0·159, and the atomic positions are known to 0·1 Å accuracy in internal segments of the molecule.The iron atom lies 0·22(3) Å from the plane of the porphyrin, 0·25 Å closer than in deoxymyoglobin, and the F helix has moved by a similar amount. Oxygen binds to the iron in a bent, end-on arrangement, with FeOO = 115(5) ° and FeO = 1·83(6) Å. The mean FeN(porphyrin) bond length is 1·95(6) Å, 0·08 Å shorter than in deoxymyoglobin, but the difference is not significant compared to the experimental error. FeN_ε(His8F) is 2·07(6) Å, the same as in model compounds. Movements of the haem, iron, F helix and FG corner on oxygenation are similar to those found in the T-R state transition in haemoglobin, but are smaller in magnitude.     

Caffeine complexes of platinum(II): crystal structure of cis-[Pt(C8H10N4O2)2Cl2]·0.4H2O

The preparations are reported of cis[Pt(caffeine)₂Cl₂]·0.4H₂O, Pd(caffeine)₂Cl₂, the methanol adduct of the previously known compound K[Pt(caffeine)Cl₃], and Pt(caffeine)(cytidine)Cl₂. Crystals of [Pt(caffeine)₂Cl₂]·0.4H₂O are tetragonal P4₂/n with a = 13.156(2) 0?, c = 12.734(2) 0?, Z = 4. The structure was refined on 945 reflections to R = 0.025. The molecule is cis with a crystallographic two-fold axis bisecting the ClPtCl and NPtN angles. The Pt is square planar with PtN and PtCl bonds of 2.029(5) Å and 2.271(2) Å respectively. There is a 5.4° dihedral angle between the imidazole and pyrimidine rings, and the imidazole ring is rotated away from the coordination plane by 86.4°. Symmetry related caffeine units pack parallel to each other with an inter-ring separation of 3.45 Å.     

Preparation and crystal structure of bis(acetato)(trans-1,2-diaminocyclohexane)platinum(II)

The structure of the complex [Pt(trans-1,2-di- aminocyclohexane) (acetate)₂]·H₂O has been determined by X-ray diffraction. This racemic compound is orthorhombic, space group Aba2, a = 20.813(9), b = 7.926(5), c = 17.296(8) Å, Z = 8. The structure was refined on 1214 nonzero Cu Kα reflections to R = 0.028. The square planar environment of Pt includes the amino groups of the diamine in cis positions and oxygens from two monodentate acetates. The PtN and PtO distances average 2.00(3) and 2.02(3) Å, respectively. The bite of the diamine ligand imposes a NPtN angle of 85(1)°, whereas the small OPtO angle of 85(1)° probably results from packing effects. The average plane through the puckered cyclohexyl ring makes an angle of 19° with the PtN₂O₂ plane. The molecules are stacked by pairs along the b axis. The two molecules of each pair are 180° apart about the stacking axis, and form altogether four NH···O hydrogen bonds.     

Synthesis,structural characterization,and properties of the organothorium alkylthiolate complex [(CH3)5C5]2Th(SCH2CH2CH3)2

This contribution reports the synthesis and characterization of the organothorium alkylthiolate complex [(CH₃)₅C₅]₂Th(SCH₂CH₂CH₃)₂. This compound crystallizes in the monoclinic space group C2/c (#15) with four molecules in a cell of dimensions a=19.066(2), b=11.603(1), c=16.379(2) Å, and β=130.08(1)°. Least-squares refinement led to a value for the conventional R index (on F_o) of 0.040 for 132 variables and 2030 observations having F_o²⩾3σ(F_o²). The molecular structure consists of an unexceptional ‘bent sandwich’ [(CH₃)₅C₅]₂Th fragment coordinated to two n-propylthiolate ligands. The ThS bond distance is 2.718(3) Å; the SC(α) distance, 1.78(2) Å; the ThSC(α) angle, 108.3(5)°; and the SThS′ angle, 102.5(2)°. Contrasts are drawn with the structures of analogous actinide alkoxides     

6-Azauridine, a nucleoside with unusual ribose conformation. The molecular and crystal structure

The cytostatic analogue ribo-6-azauridine crystallizes in the orthorhombic space group P2₁2₁2₁ with eight molecules per unit cell of dimensions $\text{a = 20.230, b = 7.709, c = 12.863}\text{A}\text{?}$. A trial structure was obtained by direct methods. Least-squares refinement of co-ordinates and anisotropic thermal parameters based on 1998 reflections measured on a four-circle diffractometer led to a discrepancy index R = 4.0%. Like uridine, 6-azauridine has the anti conformation about the glycosidic bond and a C(3′)-endo sugar pucker. Unlike uridine, it exhibits a close approach of N(6) to C(2′) at only 2.814 and 2.844 Å in the two independent molecules, and a C(5′)(5′) bond that is gauche to C(4′)O(1′) but trans to C(4′)C(3′); this conformation about a C(4′)C(5′) bond has never been observed before for C(3′)-endo puckered riboses in the crystalline state. The crystal structure displays a pseudo-A face centering and very similar conformational parameters for the two independent molecules. Every OH and NH group in the structure serves as a proton donor in a hydrogen bond, including an unusual N(3)—H(3) … O(1′) link. Molecular orbital calculations by the extended Hückel method indicate that from uridine to 6-azauridine the net charge changes sign at ring positions 5 and 6 and disappears at 1.     

Crystal structure of methyl 2,6-dichloro-2,6-dideoxy-3,4-O-isopropylidene-α-D-altropyranoside. A pyranoside system close to a skew-boat conformation

The crystal structure of methyl 2,6-dichloro-2,6-dideoxy-3,4-O-isopropylidene-α-D-altropyranoside (1) has been determined by X-ray diffraction. The compound crystallizes in the orthorhombic system, space group P2₁2₁2₁, with unit-cell dimensions a  7.932, b  8.133, and c  20.447 Å. The structure was solved by the heavy-atom method and refined by the least-squares technique to an R value of 0.047 by using 736 intensities measured on a diffractometer. The pyranoside ring is close to a skew-boat conformation, with C-2 and C-5 being maximally displaced from the least-squares plane through the remaining four atoms. The H-1H-2 dihedral angle of  158° is in agreement with the J_1,2 value of 4.5 Hz. Thus the solid-state conformation appears to correspond with the conformation in solution. The dioxolane ring is in a twist form, with O-4 and, C-8 puckered on opposite sides of the plane of the other ring atoms. The pyranose-ring substituents are in equatorial and pseudoequatorial orientations. The hydrogen atoms at C-3 and C-4 are in a cis arrangement. The orientations of both the methoxyl group and the chloromethyl group with respect to the ring are gauche—trans. The exocyclic anomeric C-1O-1 bond-distance (1.39 Å) is the shortest CO bond in the structure. The intracyclic CO bonds are significantly different, C-1O-5 being less than C-5O-5.     

MonodentateN coordination of 1aza4oxo1,3butadienes [R1NC(R2)C(R3)O] to platinum(II). X-ray structure of trans-[PtCl2(PEt3){σ-N-(t-BuNCHC(Me)O)}]

Reaction of dimeric trans-[PtCl₂(PR₃)]₂ with 1-aza-4-oxo-1,3-butadienes [R¹NC(R²)C(R³)O, R³ = Me, Ph, OMe, NEt₂] in a 1:2 molar ratio results in almost quantitative formation of mononuclear complexes trans·[PtCl₂(PR₃){σ-N-(R¹NC(R²)C(R³)O)}]. The ligands are bonded in the monodentate σ-N bonding mode to the platinum(II) centre. This has been established by an X-ray structure determination of trans-[PtCl₂(PEt₃){σ-N-(t-BuNCHC(Me)O)}]. Crystals of the latter compound are orthorhombic with space group Pc2₁n; cell constants are a = 14.712(3), b = 15.053(2), c = 9.025(5) Å, Z= 4 and R_w = 0.056 for 3281 reflections. The 1aza4oxol,3butadiene (α-iminoketone for R³ is alkyl or aryl) has the E-configuration about the imine bond (CN 1.34(4) Å), with a C(5)C(6) distance of 1.44(5) Å and a NC(5)/ C(6)O torsion angle of 89(4)°. As a result of this ligand conformation, the acetyl hydrogen atoms are positioned (on average) into the neighbourhood of the Pt-atom above the Pt-coordination plane. Infrared and NMR (¹H, ¹³C, ³¹p) data show that these structural features are also predominant in solution.     

Synthesis and spectroscopic properties of CpRuCl(R-DAB(4e)) and CpRuCo(CO)3(R-DAB(6e)). Part XVII. X-ray structure determination of CpRuCo(CO)3(t-Bu-DAB(6e))

CpRuCl(PPh₃)₂ reacted with excess R-DAB in refluxing toluene to give CpRuCl(R-DAB(4e)) (1a: R = i-Pr; 1b: R = t-Bu; 1c: R = neo-Pent; 1d: R =p-Tol). ¹H NMR and ¹³C NMR spectroscopic data indicated that in these complexes the R-DAB ligand is bonded in a chelating 4e coordination mode.Reaction of 1a and 1b with one equivalent of [Co(CO)₄]⁻ afforded CpRuCo(CO)₃(R-DAB(6e)) (2a: R = i-Pr; 2b: R = t-Bu). The structure of 2b was determined by a single crystal X-ray structure determination. Crystals of 2b are monoclinic, space group P2₁/n, with four molecules in a unit cell of dimensions: a = 16.812(4), b = 12.233(3), c = 9.938(3) Å and β = 105.47(3)°. The structure was solved via the heavy atom method and refined to R = 0.060 and R_w = 0.065 for the 3706 observed reflections. The molecule contains a RuCo bond of 2.660(3) Å and a cyclopentadienyl group that is η⁵-coordinated to ruthenium [RuC(cyclopentadienyl) = 2.208(3) Å (mean)]. Two carbonyls are terminally coordinated to cobalt (CoC(1) = 1.746(7) and CoC(2) = 1.715(6) Å) while the third is slightly asymmetrically bridging the RuCo bond (RuC(3) = 2.025(6) and CoC(3) = 1.912(6) Å). The RuC(3)O(3) and CoC(3)O(3) angles are 138.4(5)° and 136.5(5)°, respectively. The t-Bu-DAB ligand is in the bridging 6e coordination mode: σ-N coordinated to Ru (RuN(2) = 2.125(4) Å), μ₂-N′ bridging the RuCo bond and η²-CN coordinated to Co (RuN(1) = 2.113(5), CoN(1) = 1.941(4) and CoC(4) = 2.084(5) Å). The η²-CN′ bonded imine group has a bond length of 1.394(7) Å indicating substantial π-backbonding from Co into the anti-bonding orbital of this CN bond.¹H NMR spectroscopy indicated that 2a and 2b are fluxional on the NMR time scale. The fluxionality of 6e bonded R-DAB ligands is rarely observed and may be explained by the reversible interchange of the σ-N and η²-CN′ coordinated imine parts of the R-DAB ligand.     

Crystal structure of an inhibitor and a model for inhibition of replicative DNA synthesis

6-(p-Hydroxyphenylhydrazino)-uracil is an antimicrobial agent that selectively blocks replicative DNA synthesis in Bacillus subtilis by inhibiting DNA polymerase III. The drug crystallizes as a monoclinic monohydrate, space group $\text{C2}\text{c}$, with $\text{a = 23.920(6)}\text{A}\text{̊}\text{, b = 5.587(3)}\text{A}\text{̊}\text{, c = 17.466(5)}\text{A}\text{̊}\text{, β = 101.45(8) °}$, and eight hydrated molecules per cell. Three-dimensional X-ray diffraction data were collected. The structure was solved by Patterson methods and refined to an R value of 6.8% for the 1651 data. The geometry of the uracil ring is unusual. The bond distances suggest that a resonance form involving a positively charged hydrazino nitrogen and a negatively charged carbonyl oxygen, O(4), makes a large contribution to the valence bond structure of this compound. The exocyclic C(6)N bond is short (1.335 Å), the C(6)C(5) bond distance is 1.371 Å, which is longer than in uracil, and the C(5)C(4) distance (1.396 Å) is short. The uracil ring, the linked hydrazino nitrogen, and the hydrogen on this nitrogen are in the same plane. Each uracil group is hydrogen bonded to a nearly coplanar uracil across a center of symmetry. The water molecule is also near the plane of these paired bases and forms a hydrogen bond with the uracil-linked hydrazino NH group. This paired base arrangement and the restricted rotation about the exocyclic C(6)N link that constrains the hydrazino NH group to lie near the uracil plane suggest a model for the interaction of the drug with template-primer DNA. The drug acts when cytosine is the base to be copied in the template strand, and the drug is competitive with dGTP. Both cytosine and guanine can be accommodated with little distortion of the crystal structure geometry in a manner compatible with the known geometry of DNA. The structural and biochemical aspects of the model for drug action are discussed.     

Emission Mössbauer studies of some organocobalamins

Subtle changes in Mössbauer parameters are observed while going from methyl- to ethyl- to adenosylcobalamin, and also when the ‘base’ is detached from the cobalt. The observation of these changes demonstrates that the Co-C bond, among others, remains intact after the Auger event, accompanying the electron-capture decay of the cobalt-57.The differences between ethylcobalamin and the other two organocobalamins in the magnitude of the quadrupole splittings have been interpreted on the basis of the σ-donating tendency of the organic moiety and the CoC bond length. The latter is presumably determined by the steric hindrance offered to the group in approaching the cobalt atom.The ethyl- and adenosylcobalamins in their ‘base-off’ form exhibit a larger quadrupole splitting than the corresponding ‘base-on’ form. In the ‘base-off’ form, the cobalt atom is perhaps raised above the mean plane of the four equatorial nitrogen atoms of the corrin ring, which may result in the diminution of the delocalization of the 3d_π electron density. The higher population of d_π orbitals and the enhanced metallic character of the d_z2, resulting from shrink-age of the CoC bond length, enhances the magnitude of the quadrupole splitting.     

X-ray structure of a mono(1-methylthyminato) complex of cisplatin,chloro-(1-methylthyminato-N3)-cis-diammineplatinum(II) monohydrate

The crystal structure of chloro-(1-methyltyminato- N3)-cis-diammineplatinum(II) monohydrate, cis- (NH₃)₂Pt(C₆H₇N₂O₂)Cl·H₂O, is reported. The compound crystallizes in space group P$\text{1}$ with a = 6.911(2) Å, b = 8.598(3) Å, c = 11.464(4) Å, α = 100.13(3)°, β = 120.03(3)°, γ = 93.16(3)°, Z = 2. The structure was refined to R = 0.048 and R_w = 0.057. The compound contains the deprotonated 1-methylthymine ligand coordinated to Pt through N3 (1.973(10) Å). This distance represents the shortest Pt-N3(pyrimidine-2.4-dione) bond reported so far. The two PtNH₃ bond lengths differ significantly: PtNH₃ (trans to Cl) is longer (2.052(10) Å) than PtNH₃ (trans to N3 of 1-MeT) (2.002(11) Å). The PtCl distance (2.326(3) Å) is normal, as is the large dihedral angle between the Pt coordination plane and the nucleobase (76.5°).     

Quercetin and daidzein β-apo-14’-carotenoic acid esters as membrane antioxidants

Abstract

Esterification by β-apo-14’-carotenoic acid was found to have opposite effects on antioxidant activity of quercetin (at B4’, B3’ hydroxyl) as of daidzein (at A7 hydroxyl) in phosphatidylcholine liposomes. The daidzein ester had increased activity, while quercetin had a significant decreased activity. Quantum mechanical calculations using density function theory (DFT) indicate a modest decrease in bond dissociation enthalpy, BDE, for (weakest) hydrogen–oxygen phenolic bond in daidzein from 368.4 kJ·mol^{? 1} to 367.7 kJ·mol^{? 1} compared to a significant increase in quercetin from 329.5 kJ·mol^{? 1} to 356.6 kJ·mol^{? 1} upon derivatization. These opposite changes in tendency for hydrogen atom transfer from phenolic groups to lipid radicals combined with an increase in A-to-B dihedral angle from 0.0° to 36.4° and in dipole moment from 0.40 D to 6.01 D for quercetin upon derivatization, while less significant for daidzein (36.4°–36.7° and 3.26 D–7.87 D, respectively), together provide a rationale for the opposite effect of esterification on antioxidation.     

Reverse NaCa exchange requires internal Ca and/or ATP in squid axons

Classical NaCa exchange models are based on a symmetric carrier system where Na and Ca competing from the same site, can produce net movement of the other against its electrochemical gradient. We have explored this symmetric assumption by studying the Ca_o and Na_o-dependent Na efflux in dialyzed squid axons in which proper control of both external and internal medium was achieved. The results show: (1) In axons dialyzed without Ca_i and ATP, Ca_o-dependent Na efflux cannot be detected even in the absence of Na_o. Under these conditions, the level of Na efflux (1 pmol · cm⁻² · s⁻¹) is close to that predicted by an electrical ‘leak’. (2) In axons dialyzed with Ca_i (100 μM) and without ATP, Na efflux measured in 440 mM Na_o, is about 4–5 pmol · cm⁻² · s⁻¹ and rather insensitive to Ca_o between 0 and 10 mM. However, in the absence of Na_o, a Ca_o-dependent Na efflux is observed similar in magnitude to that found in the presence of external Na. (3) In the presence of both Ca_i and ATP, Na efflux into artificial sea-water (440 mM Na, 10 mM Ca) is 18 pmol · cm⁻² · s⁻¹. In the absence of Na_o the efflux of Na is 7.5 pmol · cm⁻² · s⁻¹. In the absence of both Na_o and Ca_o the efflux is close to ‘leak’. With full Na_o but no Ca_o, the Na efflux average 12.6 pmol · cm⁻² · s⁻¹. These results indicate a marked asymmetry in the modus operandi of the NaCa exchange system with respect to Ca_i and ATP. These two substrates are required from the cis side to promote Ca_o-dependent Na efflux (reversal NaCa exchange).     

Effects of dual infections of Puccinia hordei and Erysiphe graminis on barley, cv. Zephyr

Seed tubers of the varieties King Edward, Majestic and Pentland Crown selected as ‘clean’ (lesion-free), moderately, or severely affected by gangrene lesions were planted in field experiments. Infection delayed plant emergence, increased the number of stems/plant, sometimes caused gaps in crops and was associated with increased blackleg. On average severely affected seed yielded 20% less than ‘clean’ seed. Seed infection also increased the proportion of tubers in smaller size grades so that crops from severely infected King Edward seed averaged 1·4 ton/acre (3·5 t/ha) less small ware and 2·5 ton/acre (6·3 t/ha) less large ware than ‘clean’ seed. With Majestic, small ware was increased (0·7 ton/acre (1·8 t/ha)) and large ware decreased (4·4 ton/acre(11·0 t/ha)); Pentland Crown was similarly affected (small ware increased 0·8 ton/acre (2·0 t/ha); large ware decreased 3·9 ton/acre (9·8 t/ha)). In eight of twelve experiments unselected diseased stocks yielded significantly less than ‘clean’ tubers. Other experiments compared seed stocks with different proportions of gangrene-infected seed tubers. Yields decreased as the proportion of diseased seed tubers increased, but differences were significant only when more than 60% were affected. Surprisingly, yields from ‘clean’ tubers also decreased as the proportion of diseased tubers increased in the stocks from which they were selected. Gangrene on progeny tubers after storage was not always related to the amount of gangrene visible on the seed. It was increased by riddling or wounding and decreased by dipping tubers in organo-mercury fungicide before or soon after wounding.     

Crystal and molecular structure of bis(tricyclohexylphosphine)silver(I)nitrate and bis(tricyclohexylphosphine)silver(I)perchlorate; correlation of the structures with the silver-phosphorus coupling constants

Two (1:2) silver monophosphine complexes have been studied by X-ray diffraction methods and in solution by P NMR spectroscopy. Both are monomeric and tricoordinated in the solid state but one of them, the perchlorate compound, is probably associated as a dimer species in solution from the lower ¹J(¹⁰⁷Ag³¹P) value when compared to the nitrate analogue. Previous structural correlations found in other silver-phosphine complexes have been confirmed for these new compounds. Thus, larger PAgP bond angles are associated with shorter AgP bond distances, longer Aganion bond distances and lower Lewis basicity of the anions. Selected structural data are: PAgP bond angle of 139.04(9)°, AgP bond lengths of 2.440(3) and 2.445(3) Å for the nitrate complex and 147.34(3)°, 2.429(1) Å and 2.432(1) Å, for the perchlorate one. J(¹⁰⁷Ag³¹P) is 457 Hz and 447 Hz, respectively. The complexes are triclinic, Z = 2, with the parameters: a = 9.258(2), b = 9.828(2), c = 23.385(5) Å, α = 94.73(2)°, β = 96.35(2)°, γ = 116.42(1)° (nitrate) and a = 9.505(2), b = 9.790(2), c = 23.667(6) Å, α= 99.03(2) β = 95.44(2) γ = 115.97(1)° (perchlorate).     

Adducts from mercury(I) and mercury(II) compounds with Bispyrazolylalkanes.X-Ray crystal structure of Bis(3,5-dimethylpyrazol-l-yl)methane(dicyano)-mercury(II)

The 1:1 adducts between thebis(3,5-dimethylpyrazol-1-yl)methane (L′-L′) or 2,2′-bis(pyrazol- 1-yl)propane (L″L″) ligand and HgX₂ (with X = Cl, CN or CO₂CF₃) have been obtained as well as [(L′L′)₂]Hg(ClO₄)₂ and the mercury(I) derivative (ligand)₂Hg₂(ClO₄)₂. The adducts have been characterized from analytical and spectral data (IR, proton and ¹³C NMR). Four-coordinated mercury is present in (L′L′)Hg(CN)₂, in which the metal-(NN)₂C ring adopts an asymmetric boat form. The molecular parameters are significantly different for the two independent molecules, the CHgC angles and the two Hg-N distances being 163.1(9)°and 2.55(1) plus 2.70(1) Å in the one case, and 148.2(8)° and 2.40(1) plus 2.51(1) Å, in the other; correspondingly the N-Hg-N angle, the ‘bite’ of the ligand, ranges from 79.0(5)° to 71.7(4)°, a value outside the range previously reported.     

A density functional theory study on the hydrogen bonding interactions between luteolin and ethanol

Ethanol is one of the most commonly used solvents to extract flavonoids from propolis. Hydrogen bonding interactions play an important role in the properties of liquid system. The main objective of the work is to study the hydrogen bonding interactions between flavonoid and ethanol. Luteolin is a very common flavonoid that has been found in different geographical and botanical propolis. In this work, it was selected as the representative flavonoid to do detailed research. The study was performed from a theoretical perspective using density functional theory (DFT) method. After careful optimization, there exist nine optimized geometries for the luteolin ? CH₃CH₂OH complex. The binding distance of X ? H···O, and the bond length, vibrational frequency, and electron density changes of X ? H all indicate the formation of the hydrogen bond in the optimized geometries. In the optimized geometries, it is found that: (1) except for the H2’, H5’, and H6’, CH₃CH₂OH has formed hydrogen bonds with all the hydrogen and oxygen atoms in luteolin. The hydrogen atoms in the hydroxyl groups of luteolin form the strongest hydrogen bonds with CH₃CH₂OH; (2) all of the hydrogen bonds are closed-shell interactions; (3) the strongest hydrogen bond is the O3’ ? H3’···O in structure A, while the weakest one is the C3 ? H3···O in structure E; (4) the hydrogen bonds of O3’ ? H3’···O, O ? H···O4, O ? H···O3’ and O ? H···O7 are medium strength and covalent dominant in nature. While the other hydrogen bonds are weak strength and possess a dominant character of the electrostatic interactions in nature.     

Evaluating the deterioration of peanut seeds contaminated with aflatoxin

Although aflatoxin (AFT) effects on Arachis hypogaea seed germination have been examined, there have been few, if any, investigations correlating germination capacity to field AFT levels for naturally contaminated peanuts. Here, we report this analysis for selected Federal samples containing 0–20 ppb AFT collected at intervals during both September and October 1981. Following the removal of both decoated and split seeds for screening, seeds were germinated for 5 days when both dead and severely infested seeds were discarded. The remaining seeds, normal (uninfested) ‘sprouted’, ‘sprouted’ with some infestation, ‘sprouted’ with morphological abnormalities and normal unsprouted, were both measured and transferred to fresh towels for germination and growth of seedlings. The hypocotyl and primary root lengths of the seedlings were measured. Whereas these lengths ranged from 58·0 ± 6·2 to 68·8 ± 5·6 mm for contaminated seedlings (control 72·5 ± 5·9) at 5 days, hypocotyl and root lengths were 210·0 ± 12·5–225·0 ± 23·0 for contaminated seedlings (control 226·7 ± 21·3) by day 12. Then, percentage germination was 75·0 ± 3·0−84·4 ± 2·5 for toxin seeds control 85·6 ± 1·8). These results suggest that neither the percentage germination nor hypocotyl/ root lengths could be strongly correlated with AFT levels (from natural contamination) for the selected seeds.     

The three-dimensional solution structure of mini-M conotoxin BtIIIA reveals a disconnection between disulfide connectivity and peptide fold

Conotoxins are bioactive peptides from the venoms of marine snails and have been divided into several superfamilies based on homologies in their precursor sequences. The M-superfamily conotoxins can be further divided into five branches based on the number of residues in the third loop of the peptide sequence. Recently two M-1 branch conotoxins (tx3a and mr3e) with a C1–C5, C2–C4, C3–C6 disulfide connectivity and one M-2 branch conotoxin (mr3a) with a C1–C6, C2–C4, C3–C5 disulfide connectivity were described. Here we report the disulfide connectivity, chemical synthesis and the three-dimensional NMR structure of the novel 14-residue conotoxin BtIIIA, extracted from the venom of Conus betulinus. It has the same disulfide connectivity as mr3a, which puts it in the M-2 branch conotoxins but has a distinctly different structure from other M-2 branch conotoxins. 105 NOE distance restraints and seven dihedral angle restraints were used for the structure calculations. The three-dimensional structure was determined with CYANA based on torsion angle dynamics and refinement in a water solvent box was carried out with CNS. Fifty structures were calculated and the 20 lowest energy structures superimposed with a RMSD of 0.49 ± 0.16 Å. Even though it has the M-2 branch disulfide connectivity, BtIIIA was found to have a ‘flying bird’ backbone motif depiction that is found in the M-1 branch conotoxin mr3e. This study shows that conotoxins with the same cysteine framework can have different disulfide connectivities and different peptide folds.     

Crystallographic studies of drug-nucleic acid interactions: Proflavine intercalation between the non-complementary base-pairs of cytidilyl-3′,5′-adenosine

In order to get insights into the binding of dyes and mutagens with denatured and single-stranded nucleic acids and the possible implications in frameshift mutagenesis, a 1:1 complex between the non-self-complementary dinucleoside monophosphate cytidilyl-3′,5′-adenosine (CpA) and proflavine was crystallized. The crystals belong to the tetragonal space group P4₂2₁2 with cell constants $\text{a = b = 19.38(1)}\text{A}\text{?}\text{and}\text{c = 27.10(1)}\text{A}\text{?}$. The asymmetric unit contains one CpA, one proflavine and nine water molecules by weight. The structure was determined using Patterson and direct methods and refined to an R-value of 11% using 2454 diffractometer intensities.The non-self-complementary dinucleoside monophosphate CpA forms a selfpaired parallel chain dimer with a proflavine molecule intercalated between the protonated cytosine-cytosine (C · C) pair and the neutral adenine-adenine (A · A) pair. The dimer complex exhibits a right-handed helical twist and an irregular girth. The neutral A · A pair is doubly hydrogen-bonded through the N(6) and N(7) sites (C(1′)C(1′) distance: 10.97(2) Å) and the protonated C · C pair is triply hydrogen-bonded with a proton shared between the N(3) sites (C(1′)C(1′) distance: 9.59(2) Å). To accommodate the intercalating dye, the sugars of successive nucleotide residues adopt the two fundamental conformations (5′ end: 3′-endo, 3′ end: 2′-endo), the backbone adopts torsion angle values that fluctuate within their preferred conformational domains: the PO bonds (ω, ω′) adopt the characteristic helical (gauche^?-gauche^?) conformation, the CO bonds (φ, φ′) are both in the trans domain and the C(4′)C(5′) bonds (ψ) are in the gauche⁺ region. The bases of both residues are disposed in the preferred anti domain with the glycosyl torsion angles (χ) correlated to the puckering mode of the sugar so that the cytidine residue is C(3′)-endo, low χ (12 dg), and the adenosine residue is C(2′)-endo, high χ (84 °). The intercalated proflavine stacks more extensively with the C · C pair than the A · A pair. Between 4₂-related CpA proflavine units there is a second proflavine which stacks well with both the A · A and the C · C pairs sandwiching it. Both proflavine molecules are positionally disordered. In each of its two disordered sites, the intercalated proflavine forms hydrogen-bonded interactions with only one sugar-phosphate backbone. A total of 26 water sites has been characterized of which only two are fully occupied. These hydration sites are involved in an intricate network of hydrogen bonds with both the dye and CpA and provide insights on the various modes of interactions between water molecules and between water molecules and nucleic acids.The structure of the proflavine-CpA complex shows that intercalation of planar drugs can occur between non-complementary base-pairs. This result can be relevant for understanding the strong binding of acridine dyes to denatured DNA, single-stranded RNA, and single-stranded polynucleotides. Also, the ability of proflayine to promote self-pairs of adenine and cytosine bases could provide a chemical basis for an alternative mechanism of frameshift mutagenesis.