Preparation and stability of the helical form of poly 2'-O-ethyluridylic acid

2′ (3′)-O-ethyl-CMP was prepared by alkylation of CMP with diethylsulphate in alkaline medium and deaminated to give 2′(3′)-O-ethyl-UMP, which was phosphorylated to 2′(3′)-O-ethyl-UDP. About 90% of the product consisted of the 2′ isomer. The 2′(3′)-O-ethyl-UDP was readily polymerized by $\text{E}$. $\text{coli}$ polynucleotide phosphorylase in the presence of Mn⁺⁺, but not Mg⁺⁺. The 3′-isomer did not seriously interfere with polymerization nor did it act as a chain terminator. The resulting poly 2′-O-ethyluridylic acid formed a helical structure with a stability much higher then that of poly (rU) or poly 2′-O-methyluridylic acid. It also complexed readily with poly (rA). Implications with regard to the role of the 2′-hydroxyl in nucleic acid conformation are discussed.     

Bilayers of dipalmitoyl-3-sn-phosphatidylcholine: Conformational differences between the fatty acyl chains

$\text{Dipalmitoyl}\text{-3-sn-}\text{phosphatidylcholine}$ is specifically deuterated at the C-2 position of the fatty acyl chains. Using deuterium magnetic resonance it is then possible to probe the chain conformation in the vicinity the polar head groups. Three separate quadrupole splittings are observed for bilayers of 1,2[2′-²H₂] $\text{palmitoyl}\text{-3-sn-}\text{phosphatidylcholine}$, indicating that the two chains behave differently. The synthesis of phosphatidylcholines each deuterated in only one chain allows the assignment of the three resonances. It is concluded that the beginnings of the two chains have orientations parallel and perpendicular to the bilayer normal. The data further suggest the possibility of two long-lived conformations of the glycerol constituent.     

Crystal and molecular structure of [Cu(5'-inosine monophosphate) (dipyridylamine) (H2O)]2.4H2O.

The ternary complex [Cu(5′-IMP)(dpa)(H₂O)]₂ has been prepared and its structure analyzed by x-ray diffraction. It has a dimeric structure in which the 5′-IMP ligands coordinate solely through their phosphate groups. This geometry is in marked contrast to that of another $\text{Cu}\text{5′-}\text{IMP}$ ternary complex, [Cu(5′-IMPH)(bipy)(H₂O)₂]⁺, which shows metal binding through the purine base rather than the phosphate group.     

Molecular and crystal structure of the regenerated form of (1→3)-α-d-glucan

The crystal structure of a regenerated form of (1→3)-α-d-glucan, obtained by solid state deacetylation of the triacetate derivative, has been determined by combined X-ray diffraction analysis and stereochemical model refinement. The structure crystallizes in an orthorhombic unit cell with parameters $\text{a = 16.46}\text{A}\text{?}\text{, b = 9.55}\text{A}\text{?}$ and c (fibre repeat)=8.44 Å, and space group P2₁2₁2₁. The chain conformation is nearly completely extended and is very close to a 2/1 helix, even though the dimer residue is the crystallographic repeat unit. An intramolecular O(2)  O(4)′ hydrogen bond stabilizes the conformation and extensive intermolecular hydrogen-bonding abilizes the packing. The resulting structure is sheet-like, with an alternating polarity of chain directions within the sheet. In its sheet-like character, extensive hydrogen-bonding, and insolubility in water, this polymorph of (1→3)-α-d-glucan resembles regenerated cellulose. The reliability of the structure analysis is indicated by the X-ray residual R=0.206.     

Circular dichroism study of 2'-5' dinucleoside monophosphates modified with N-2-acetylaminofluorene.

The conformational properties of four 2′ – 5′ dinucleoside monophosphates modified with $\text{N}$-2-acetylaminofluorene have been studied by circular dichroism spectroscopy. Covalent binding of this chemical carcinogen at the C₈ position of guanosine in the 2′ – 5′ dinucleoside monophosphates induces striking changes in their circular dichroic spectra depending on their base sequence and composition. The changes in CD spectra, redshift of the extrema and change of their polarity, not observed in the spectra of corresponding 3′ – 5′ derivatives modified with $\text{N}$-2-acetylaminofluorene are correlated with the difference in the configuration of 2′ – 5′ and 3′ – 5′ dinucleoside monophosphates and discussed in respect to the intramolecular stacking interactions.     

Delineation of the intimate details of the backbone conformation of pyridine nucleotide coenzymes in aqueous solution.

The noise free 300 MHz ¹H NMR spectra of β-DPN⁺, recorded in the Fourier mode at 12° and 68°C have been completely analysed by extensive computer simulation. It is shown, whether the coenzyme exists as an equilibrium mixture of folded ? extended forms (12°C) or in overwhelminghly extended forms (68°C), the backbone of both the nicotinamide and adenine fragments preferentially exist in ${}^2\text{E-gg-g′g′}$ conformation. This orientation is significantly different from those reported in the solid state for the extended species in contact with the enzyme where ${}^2\text{E-tg-g′g′}$ and ${}^3\text{E-tg-g′g′}$ orientations have been observed. It is suggested that specific interactions of the backbone with the various amino acid residues in the enzyme induces conformational aberrations in the backbone. Intimate details of the backbone conformation of the extended forms of AcPy-DPN⁺ and β-TPN⁺ are also presented.     

One predominant 5′-undecanucleotide in adenovirus 2 late messenger RNAs

Oligonucleotides containing the 5′ termini of adenovirus 2 mRNA are selectively retained on columns of dihydroxyboryl cellulose. When total late adenovirus 2 mRNA was treated with RNAase T1, a single 5′ terminal oligonucleotide was isolated, although in several states of methylation. This oligonucleotide has the general structure m⁷GS^5′ppp^5′$\text{A}$mCmU(C₄,U₃)G. Since at least twelve individual species of mRNA must be present late after infection, this finding was unexpected and its significance is discussed.     

Structural investigations of mode of action of drugs. I. Molecular structure of mitomycin C.

The crystal and molecular structure of the antitumor antibiotic mitomycin C has been determined by X-ray diffraction. The space group is monoclinic P2₁ with cell dimensions $\text{a}\text{=77.988(3)}$, $\text{b}\text{=20.355(7)}$, $\text{c}\text{=9.679(3)}\text{A}\text{?}$, β=95.99°(1) and Z=4. The structure was solved by direct methods. There are two independent molecules per asymmetric unit. The structure was refined to an R value of 0.049 for 2151 observed reflections measured on diffractometer. The benzoquinone ring is slightly deviated from planarity. The N4 in the indole ring behaves like an amide nitrogen owing to its participation in the conjugated benzoquinoid system. The five membered ring through C1, C2, C3, N4 and C9a adopts an envelope conformation. The molecules are held together in crystal by the hydrogen bonds. All nitrogens except N4 are involved in the hydrogen bonding. Studies with models of drug and DNA indicate two different kinds of mechanism for crosslinking.     

Neutron diffraction studies on phosphatidylcholine model membranes. I. Head group conformation

Neutron diffraction experiments on selectively deuterated lipids provide a new method of determining to a segmental resolution the mean conformation of a lipid molecule as projected along the bilayer normal, despite the high amount of disorder that exists in these bilayers. In addition, a time-averaged picture of the extent of the positional fluctuations of the individual segments in this direction can be given. This is demonstrated for a multilamellar system of bilayers of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine. In this paper the head group region of the molecule is examined and this carries the zwitterionic phosphocholine group that determines the electrostatic interaction in the bilayer. Samples deuterated at four different positions in the head group region were measured as oriented samples at 6% ($\text{w}\text{w}$) water content at 20 °C (L_β′ phase) and at 10% ($\text{w}\text{w}$) at 70 °C (L_α phase) and as unsonicated dispersions with 25% ($\text{w}\text{w}$) water at 28 °C (L_β′ phase) and 50 °C (L_α phase). From the oriented samples, reflections up to ten orders, and from the powder type samples only four orders, were collected. The derived structure factors for the deuterated segments were fitted assuming a Gaussian distribution of the segments along the bilayer normal. The mean label position was determined for each label under different conditions of water content and temperature with a precision of better than ± 1 ångström in most cases. The data clearly show that the average orientation of the zwitterionic phosphocholine group is almost parallel to the membrane surface in the gel state (L_β′) as well as in the liquid crystalline state (L_α). It is interesting to note that in a recent dielectric investigation on this multilamellar system at 25% ($\text{w}\text{w}$) water content the same mean orientation of the dipole was found (Shepherd &; Büldt, 1978).     

Solution conformation and crystal structure of methyl 3,6-dideoxy-β-d-ribohexopyranoside,an immunodominant sugar of O-antigens

The crystal structure of methyl 3,6-dideoxy-β-d-ribohexopyranoside monohydrate was determined by direct methods. Crystals are monoclinic, space group P2₁, with cell dimensions $\text{a = 9.089(1), b = 7.668(1), c = 6.956(1)}\text{A}\text{?}\text{, β = 101.12°}$. The molecule adopts the ⁴C₁ chair conformation. The same conformation was also found in both aqueous and chloroform solutions. The pyranose ring is only slightly distorted, and the consequences of this observation on antigen structure are discussed.     

Analysis of the inhibitor binding to the nucleoside site of phosphorylase b

The binding of inhibitors to site I of rabbit muscle phsphorylase $\text{b}$ has beenstudied kinetically and thermodynamically for caffeine, adenine and adenosine. The effect of ligands on the tertiary structure has been investigated by studying the protection against 5,5′-dithiobis (2-nitrobenzoic acid) (DTNB) titration of the slow-reacting sulphydryl groups of the enzyme. Calorimetric and cysteinyl protection data taken together suggest that these inhibitors bind to both sites N and I even under conditions of saturation by glucose. Calorimetric results show that inhibitor binding to sites I and N at 25°C is driven enthalpically, although both $\text{Δ}\text{H}$ and $\text{Δ}\text{S}$ of interaction are significant. We conclude that attractive dispersion forces ought to be the main ones responsible for inhibitor binding to site I. AMP-activated phosphorylase $\text{b}$ is inhibited by both caffeine and adenine by cooperative and exclusive binding to the inactive $\text{T}$ conformation. The binding of the substrate (phosphate) and AMP when adenine is present was found to be exlusive to the active $\text{R}$ conformation, whereas non-exclusive binding of the activator was observed when caffeine was added.     

Dibutyryl cGMP: inhibitor of the effect of cholecystokinin and gastrin on the guinea pig gallbladder in vitro

N², O²-di-butyryl guanosine 3′:5′ monophosphate (Bt₂ cGMP), a known competitive and selective inhibitor of the effect of cholecystokinin on the pancreatic acinar cells $\text{in}$$\text{vitro}$ was tested for its effect on the guinea pig gallbladder $\text{in}$$\text{vitro}$. Bt₂ cGMP inhibited competitively the contractile effect of cholecystokinin octapeptide, and also inhibited the contraction induced by sulfated gastrin-17. Bt₂ cGMP failed to inhibit the contraction induced by bombesin, acetylcholine or histamine. The 8-bromo derivative of cGMP and the dibutyryl derivative of cAMP did not affect contraction stimulated by cholecystokinin octapeptide. Since it is specific for gastrincholecystokinin peptides, and not restricted to the pancreas, Bt₂ cGMP could be used to recognize the action of these peptides.     

Conformation of acetylaminofluorene and aminofluorene modified guanosine and guanosine derivatives

Acetylaminofluorene and aminofluorene modified Guo, GMP, d(GpA) and d(ApG) have been studied by circular dichroism and ¹H nuclear magnetic resonance. Aminofluorene modified Guo is preferentially in the $\text{anti}$ conformation and acetylaminofluorene modified Guo in the $\text{syn}$ conformation. It is proposed that the $\text{anti}$ conformation of aminofluorene modified Guo is stabilized by an intra molecular hydrogen bond between the NH group of aminofluorene residue and the 5′-OH group of the sugar. The results on the modified dinucleoside monophosphates are analyzed according to this hypothesis.     

Mechaism of Arthrobacter sialophilus neuraminidase: The binding of substrates and transition-state analogs

2-Deoxy-2,3-dehydro-N-acetylneuraminic acid and its methyl ester are competitive inhibitors of Arthrobacter sialophilus neuraminidase with K_i = 1.4 × 10^?6$\text{M}$ and 4.8 × 10^?5$\text{M}$, respectively. The K_m for the substrate, N-acetylneuraminlactose, is 1.0 × 10^?3$\text{M}$. These data, taken together with the conformation of these compounds, indicate that these compounds are transition-state analogs of the enzyme. These results also suggest that the substrate upon binding to neuraminidase is distorted to a conformation approaching that of a half-chair.     

Effect of specific inhibitors on mitochondrial DNA replication in HeLa cells

The crystal structure of an orthorhombic form of 2′-0-methyl cytidine was determined from three dimensional X-ray diffraction data. The two molecules in each asymmetric unit have C2-$\text{endo}$ C3-$\text{exo}$ puckered furanose rings. This differs from the C3-$\text{endo}$ puckering observed for cytidine (1) and it may have some relevance to the kinks that appear at the two 2′-0-methylated nucleotides in the anticodon phosphate ester backbone of the phe tRNA structure (2). This work and other studies (3,4) show that the presence of a 2′-0-methyl group does not prevent the furanose moiety from adopting its most commonly observed configurations. 2′-0-methyl nucleotides make up a small percentage of the residues in HnRNA, rRNA, tRNA and mRNA and therefore their conformational nuances are of interest.     

Visualization of drug--nucleic acid interactions at atomic resolution. V. Structure of two aminoacridine--dinucleoside monophosphate crystalline complexes, proflavine--5-iodocytidylyl (3'-5') guanosine and acridine orange--5-iodocytidylyl (3'-5') guanosine

Acridine orange and proflavine form complexes with the dinucleoside monophosphate, 5-iodocytidylyl(3′–5′)guanosine. The acridine orange-iodoCpG² crystals are monoclinic, space group P2₁, with unit cell dimensions $\text{a = 14.36}\text{A}\text{?}\text{, b = 19.64}\text{A}\text{?}\text{, c = 20.67}\text{A}\text{?}$, β = 102.5 °. The proflavine-iodoCpG crystals are monoclinic, space group C2, with unit cell dimensions $\text{a = 32.14}\text{A}\text{?}\text{, b = 22.23}\text{A}\text{?}\text{, c = 18.42}\text{A}\text{?}$, β = 123.3 °. Both structures have been solved to atomic resolution by Patterson and Fourier methods, and refined by full matrix least-squares.Acridine orange forms an intercalative structure with iodoCpG in much the same manner as ethidium, ellipticine and 3,5,6,8-tetramethyl-N-methyl phenanthrolinium (Jain et al., 1977, Jain et al., 1979), except that the acridine nucleus lies asymmetrically in the intercalation site. This asymmetric intercalation is accompanied by a sliding of base-pairs upon the acridine nucleus and is similar to that observed with the 9-aminoacridine-iodoCpG asymmetric intercalative binding mode described in the previous papers (Sakore et al., 1977, Sakore et al., 1979). Basepairs above and below the drug are separated by about 6.8 Å and are twisted about 10 °; this reflects the mixed sugar puckering pattern observed in the sugar-phospate chains: C3′ endo (3′–5′) C2′ endo (i.e. each cytidine residue has a C3′ endo sugar comformation, while each guanosine residue has a C2′ endo sugar conformation), alterations in glycosidic torsional angles and other small but significant conformational changes in the sugar-phosphate backbone.Proflavine, on the other hand, demonstrates symmetric intercalation with iodoCpG. Hydrogen bonds connect amino groups on proflavine with phosphate oxygen atoms on the dinucleotide. In contrast to the acridine orange structure, base-pairs above and below the intercalative proflavine molecule are twisted about 36 °. The altered magnitude of this angular twist reflects the sugar puckering pattern that is observed: C3′ endo (3′–5′) C3′ endo. Since proflavine is known to unwind DNA in much the same manner as ethidium and acridine orange (Waring, 1970), one cannot use the information from this model system to understand how proflavine binds to DNA (it is possible, for example, that hydrogen bonding observed between proflavine and iodoCpG alters the intercalative geometry in this model system).Instead, we propose a model for proflavine-DNA binding in which proflavine lies asymmetrically in the intercalation site (characterized by the C3′ endo (3′–5′) C2′ endo mixed sugar puckering pattern) and forms only one hydrogen bond to a neighboring phosphate oxygen atom. Our model for proflavine-DNA binding, therefore, is very similar to our acridine orange-DNA binding model. We will describe these models in detail in this paper.     

Synthesis and antitumor activity of 1-beta-D-arabinofuranosylcytosine conjugates of cortisol and cortisone.

Superior antitumor activity of 1-β-D-arabinofuranosylcytosine (ara-C) conjugates of prednisolone and prednisone against L1210 leukemic mice, based on ara-C content, has encouraged us to synthesize 5′-(cortisone-21-phosphoryl)-1-β-D-arabinofuranosylcytosine (I) and 5′-(cortisone-21-phosphoryl)-1-β-d-arabinofuranosylcytosine (II) by condensation of N⁴,2′,3′-triacetyl-1-β-d-arabinofuranosylcytosine 5′-monophosphate with cortisol and cortisone in the presence of N,N′-dicyclohexylcarbodiimide at room temperature followed by removing the acetyl groups in 2 N methanolic ammonia in 20% yield. The conjugates I and II inhibited the $\text{in}\text{vitro}$ growth of L1210 by 50% (ED₅₀) at 0.25 μM and 0.07 μM, respectively, while ara-C showed ED₅₀ 0.1 μM. However, the conjugates I and II exhibited 287% and 238% of $\text{T}\text{C}$ at 50 mg/kg/day × 5 doses against L1210 leukemic mice, respectively, while ara-C at 25 mg and 50 mg/kg/day × 5 gave the respective 127% and 110% of $\text{T}\text{C}$.     

Molecular conformation of prostaglandin A1

The crystal and molecular structure of prostaglandin A₁ (PGA₁) has been determined by X-ray diffraction. (Space group P2₁2₁2₁, $\text{a}\text{= 18.10}\text{A}\text{?}\text{A}$, $\text{b}\text{= 21.09}\text{A}\text{?}\text{A}\text{,}\text{c}\text{= 5.42,}\text{Z}\text{= 4)}$. Comparison of the structure of PGA₁ with that of PGF_1β (determined previously as the tribromobenzoate) indicates significant differences in the relative intramolecular side chain orientations. However, conformation and torsional angles within each side chain of PGA₁ are retained with little or no change in comparison with PGF_1β, indicating the presence of the C₁₅-bromobenzoate groups in the latter have had little effect in altering internal side chain conformation in the solid state. The overall differences in relative side chain orientation in PGA₁ are attributable to chemical and conformational changes in the substituted cyclopentane moiety (C₈-C₁₂).     

Chloride and tropolonato mixed complexes of uranium(IV) and thorium(IV)

The synthesis of some new chloride and tropolonato mixed complexes of the general composition MCl_4?nTrop_n·mL (M = U^IV, Th^IV; n = 1, 2, 3; m = $\text{1}\text{2}$, $\text{2}\text{3}$, 1, 2 and 3; L = DME, THF, LiCl and 18-crown-6) by the reaction of uranium and thorium tetrachlorides with M′Trop (M′ = Tl and Li) is described. The residual chloride atoms in UTropCl₃·THF undergo further substitution reactions involving TlCp, while UTrop₂Cl₂·THF shows a different behaviour toward TlCp, since Cp₃UCl as one of the main reaction products is obtained. Protolysis of Cp₂U(NEt₂)₂ by HTrop affords a mixture of CpUTrop₃ and Cp₂UTrop₂. Finally both UTropCl₃·THF and UCl₄ react directly with HTrop, the nature of the resulting products depending on the reaction solvent.     

Complex formation between transfer RNAs with complementary anticodons: use of matrix bound tRNA

It is shown that yeast tRNA^Phe, chemically coupled by its oxidized 3′CpCpA end behaves exactly as free tRNA^Phe in its ability to form a specific complex with $\text{E. coli}$ tRNA₂^Glu having a complementary anticodon. The results support models of tRNA in which the 3′CpCpA_OH end and the anticodon are not closely associated in the tertiary structure, and provide a convenient tool of general use to characterize others pairs of tRNA having complementary anticodons, as well as for highly selective purification of certain tRNA species.