Role of magnesium ion in the interaction between chromomycin A3 and DNA: binding of chromomycin A3-Mg2+ complexes with DNA.

Chromomycin A3 is an antitumor antibiotic which blocks macromolecular synthesis via reversible interaction with DNA template only in the presence of divalent metal ions such as Mg2+. The role of Mg2+ in this antibiotic-DNA interaction is not well understood. We approached the problem in two steps via studies on the interaction of (i) chromomycin A3 and Mg2+ and (ii) chromomycin A3-Mg2+ complex(es) and DNA. Spectroscopic techniques such as absorption, fluorescence, and CD were employed for this purpose. The results could be summed up in two parts. Absorption, fluorescence, and CD spectra of the antibiotic change upon addition of Mg2+ due to complex formation between them. Analysis of the quantitative dependence of change in absorbance of chromomycin A3 (at 440 nm) upon input concentration of Mg2+ indicates formation of two types of complexes with different stoichiometries and formation constants. Trends in change of fluorescence and CD spectroscopic features of the antibiotic in the presence of Mg2+ at different concentrations further corroborate this result. The two complexes are referred to as complex I (with 1:1 stoichiometry in terms of chromomycin A3:Mg2+) and complex II (with 2:1 stoichiometry in terms of chromomycin A3:Mg2+), respectively, in future discussions. The interactions of these complexes with calf thymus DNA were examined to check whether they bind differently to the same DNA. Evaluation of binding parameters, intrinsic binding constants, and binding stoichiometry, by means of spectrophotometric and fluorescence titrations, shows that they are different. Distinctive spectroscopic features of complexes I and II, when they are bound to DNA, also support that they bind differently to the above DNA. Measurement of thermodynamic parameters characterizing their interactions with calf thymus DNA shows that complex I-DNA interaction is exothermic, in contrast to complex II-DNA interaction, which is endothermic. This feature implies a difference in the molecular nature of the interactions between the complexes and calf thymus DNA. These observations are novel and significant to understand the antitumor property of the antibiotic. They are also discussed to provide explanations for the earlier reports that in some cases appeared to be contradictory.     

Crystal structure of Mg2+- and Ca2+-bound Gla domain of factor IX complexed with binding protein

Factor IX is an indispensable protein required in the blood coagulation cascade. It binds to the surface of phospholipid membrane by means of a gamma-carboxyglutamic acid (Gla) domain situated at the N terminus. Recently, we showed that physiological concentrations of Mg2+ ions affect the native conformation of the Gla domain and in doing so augment the biological activity of factor IXa and binding affinity with its binding protein even in the presence of Ca2+ ions. Here we report on the crystal structures of the Mg2+/Ca2+-bound and Ca2+-bound (Mg2+-free) factor IX Gla domain (IXGD1-46) in complex with its binding protein (IX-bp) at 1.55 and 1.80 A resolutions, respectively. Three Mg2+ and five Ca2+ ions were bound in the Mg2+/Ca2+-bound IXGD1-46, and the Mg2+ ions were replaced by Ca2+ ions in Mg2+-free IXGD1-46. Comparison of Mg2+/Ca2+-bound with Ca2+-bound structures of the complexes showed that Mg2+ ion, which formed a bridge between IXGD1-46 and IX-bp, forced IXGD1-46 to rotate 4 degrees relative to IX-bp and hence might be the cause of a more tight interaction between the molecules than in the case of the Mg2+-free structure. The results clearly suggest that Mg2+ ions are required to maintain native conformation and in vivo function of factor IX Gla domain during blood coagulation.     

Synthesis, characterization and DNA binding of magnesium-ciprofloxacin (cfH) complex [Mg(cf)2] * 2.5H2O

Interactions of the tested systems (title compound [Mg(cf)(2)] * 2.5H(2)O (1), ciprofloxacin (cfH) and ciprofloxacin in the mixture with MgCl(2)), with single and double stranded calf thymus DNA, poly[d(AT)] * poly[d(AT)] and poly[d(GC)] * poly[d(GC)] were studied by UV-spectrophotometric (melting curves) and fluorescence emission measurements. Pronounced quenching of ciprofloxacin's fluorescence intensity has been observed for all the tested compounds after titration with various GC containing DNA molecules. It seems probable that quenching originates in the electron transfer from guanine to the photo-excited fluoroquinolone. The UV-spectrophotometric results obtained for 1 are substantially different from the other solutions and the biggest differences were observed for GC containing DNAs. Solution of 1 provokes a large thermal destabilization of poly[d(GC)] * poly[d(GC)]. This process is irreversible which suggests that the species present in solution of 1 alone inhibit re-annealing by associating irreversibly with the single strands. We have realized that aqueous solutions of 1 are colloidal and we propose that colloidal particles are involved in specific binding to GC containing sequences, most probably in the major groove of DNA.     

Correlation of antibacterial activity of some N-[5-(2-furanyl)-2-methyl-4-oxo-4H-thieno[2,3-d]pyrimidin-3-yl]-carboxamide and 3-substituted-5-(2-furanyl)-2-methyl-3H-thieno[2,3-d]pyrimidin-4-ones with topological indices using Hansch analysis

The antimicrobial activity of the N-[5-(2-furanyl)-2-methyl-4-oxo-4H-theino[2,3-d]pyrimidin-3-y1]-carboxamides and 3-substituted-5-(2-furanyl)-2-methyl-3H-thieno[2,3-d]pyrimidin-4-ones was correlated with different topological indices using Hansch analysis. Good correlations were obtained through a simple regression equation with third order molecular connectivity index (3chi). The developed QSAR models were crossvalidated by leave-one-out technique.     

Crystal structure and magnetic measurements of [Cr(en)3] [ZnCl4]Cl

The compound [Cr(en)₃][ZnCl₄]Cl has been synthesized by reaction of CrCl₃·6H₂O, Zn and [Cr(en)₃]₂(SO₄)₃ in HCl. Its molecular and crystalline structure was determined by X-ray diffraction methods, being monoclinic, P2₁/c, a=21.215(3), b=12.532(2), c=13.707(2) Å, β=95.21°, V= 3629(2) ų, D_x=1.738g cm⁻³, MW=474.9, Z= 8, F(000)=1928, λ(Mo Kα)=0.71069 Å, μ(Mo Kα)= 27.04 cm⁻¹, 288 K. No significant exchange interactions between Cr(III) cations in the crystalline lattice were found. Curie-Weiss behavior was found in the three directions tested (g₁=2.06±0.02,g₂= 2.08±0.02,g₃=2.09±0.01), T=1.2-1.4 K.     

Calcium binding cyclic hexapeptide. Crystal structure of cyclo-(L-prolyl-glycyl)3 calcium complex

The synthetic cyclic hexapeptide (L-prolyl-glycyl)3 forms a 2:1 complex with Ca2+ ion. The cation is sandwiched between the two peptide molecules. The glycyl carbonyls from each of the peptides are octahedrally coordinated to the cation with an average calcium oxygen coordination distance of 2.26A. Both the molecules coordinating to the calcium ion have three fold symmetry, but show significant conformational differences. In one of the peptides of the sandwich, the alternate carbonyls point to the opposite sides of the peptide ring while in the other, all the six carbonyls point to the same side of the ring. Three NH ... O hydrogen bonds between the peptides add to the stability of the sandwich.     

8-[3H]Hydroxy-2-(di-n-propylamino) tetralin binding sites in goldfish retina

The binding sites of 8-[³H]hydroxy-2-(di-n-propylamino)tetralin ([³H]DPAT) were characterized in the retina of goldfish in order to evaluate the selectivity of the ligand for serotonin_1A (5HT_1A) receptors. Specificity of the binding was performed in the presence of serotonergic and dopaminergic agonists and antagonists. Buspirone, spriroxatrine and 5-methoxy-N,N-dimethyltryptamine were potent inhibitors, followed by propranolol, citalopram, imipramine and desipramine. Serotonin was not a potent inhibitor, and its interaction with the binding sites of [³H]DPAT was complex. Nomifensine displayed an important inhibition, however, other dopamine uptake blockers, such as bupropion and GBR-12909, were less potent. Haloperidol was also a good inhibitor, but the D₁ receptor agonist, SKF-38393, the D₂ receptor antagonist, sulpiride, and dopamine did not inhibit the binding. GppNHp inhibited the binding in the micromolar range. The analysis of saturation experiments by isotopic dilution, using buspirone to determine nonspecific binding, revealed two sites. The number of binding sites defined by buspirone were higher than the ones defined by nomifesine. The specific binding, using buspirone for definition, was reduced by the intraocular injection of 6-hydroxydopamine. This investigation demonstrates that [³H]DPAT labels 5HT_1A receptors in goldfish retina, but also interacts with a non-5HT receptor site. These receptors seem to be localized in dopaminergic neurons.     

Crystal structure of [Sm(OPMePh2)4I2]I. A cationic octahedral complex of samarium(III)

The crystal structure of [Sm(OPMePh₂)₄I₂]I, 1, was determined by X-ray diffraction and refined anisotropically to a final R value of 0.067 from 3040 reflections with I>3.0σ(I). The space group was P2/a and Z=2. The unit cell dimensions were: a= 17.777(6), b=13.559(2), c=11.656(4) Å, α=γ= 90.0 and β=97.25(3)°. The cation geometry was octahedral with the Sm(III) bonded to two mutually trans I⁻ ions and four OPMePh₂ groups. A third non-bonded I⁻ was present elsewhere in the cell. The SmI and SmO distances were 3.077(1) and 2.27(1) Å respectively. Two of the SmOP angles were 172.1(6)° and the other two were 162.0(6)°.     

Crystal structure of memapsin 2 (beta-secretase) in complex with an inhibitor OM00-3

The structure of the catalytic domain of human memapsin 2 bound to an inhibitor OM00-3 (Glu-Leu-Asp-LeuAla-Val-Glu-Phe, K(i) = 0.3 nM, the asterisk denotes the hydroxyethylene transition-state isostere) has been determined at 2.1 A resolution. Uniquely defined in the structure are the locations of S(3)' and S(4)' subsites, which were not identified in the previous structure of memapsin 2 in complex with the inhibitor OM99-2 (Glu-Val-Asn-LeuAla-Ala-Glu-Phe, K(i) = 1 nM). Different binding modes for the P(2) and P(4) side chains are also observed. These new structural elements are useful for the design of new inhibitors. The structural and kinetic data indicate that the replacement of the P(2)' alanine in OM99-2 with a valine in OM00-3 stabilizes the binding of P(3)' and P(4)'.     

Similar binding of the carcinostatic drugs cis-[Pt(NH3)2Cl2] and [Ru(NH3)5Cl] Cl2 to tRNAphe and a comparison with the binding of the inactive trans-[Pt(NH3)2Cl2] complex - reluctance in binding to Watson-Crick base pairs within double helix.

A comparative study of the binding of square planar cis- and trans-[Pt(NH3)2Cl2] complexes and the octahedral [Ru(NH3)5(H2O)]3+ complex to tRNAphe from yeast was carried out by X-ray crystallography. Both of the carcinostatic compounds, cis-[Pt(NH3)2Cl2] and [Ru(NH3)5(H2O)]3+ show similarities in their mode of binding to tRNA. These complexes bind specifically to the N(7) positions of guanines G15 and G18 in the dihydrouridine loop. [Ru(NH3)5(H2O)]3+ has an additional binding site at N(7) of residue G1 after extensive soaking times (58 days). A noncovalent binding site for ruthenium is also observed in the deep groove of the acceptor stem helix with shorter (25 days) soaking time. The major binding site for the inactive trans-[Pt(NH3)Cl2] complex is at the N(1) position of residue A73, with minor trans-Pt binding sites at the N(7) positions of residues Gm34, G18 and G43. The similarities in the binding modes of cis-[Pt(NH3)2Cl2] and [Ru(NH3)5(H2O)]3+ are expected to be related to their carcinostatic properties.     

Crystal structures of the dimeric cobalt compounds [Co(PPhMe2)(CO)3]2 and [Co2(PPhMe2)3(CO)5]

The molecular structures of the dimeric cobalt compounds [Co(PPhMe₂)(CO)₃]₂ and [Co₂(PPhMe₂)₃(CO)₅] have been characterised by X-ray crystallography. The compound [Co₂(PPhMe₂)₃(CO)₅] contains both bridging and terminal carbonyl groups in the solid state.     

Quantification of isomeric equilibria formed by metal ion complexes of 8-[2-(phosphonomethoxy)ethyl]-8-azaadenine (8,8aPMEA) and 9-[2-(phosphonomethoxy)ethyl]-8-azaadenine (9,8aPMEA). Derivatives of the antiviral nucleotide analogue 9-[2-(phosphonomethoxy)ethyl]adenine (PMEA)

The acidity constants of the two-fold protonated acyclic 9-[2-(phosphonomethoxy)ethyl]-8-azaadenine, H2(9,8aPMEA)(+)(-), and its 8-isomer, 8-[2-(phosphonomethoxy)ethyl]-8-azaadenine, H2(8,8aPMEA)(+)(-), both abbreviated as H2(PA)(+)(-), as well as the stability constants of their M(H;PA)+ and M(PA) complexes with the metal ions M2+=Mg2+, Ca2+, Sr2+, Ba2+, Mn2+, Co2+, Ni2+, Cu2+, Zn2+ or Cd2+, have been determined by potentiometric pH titrations in aqueous solution at I=0.1 M (NaNO3) and 25 degrees C. Application of previously determined straight-line plots of log K(M)M(R-PO3) versus pK(H)H(R-PO3)for simple phosph(on)ate ligands, R-PO3(2-), where R represents a residue without an affinity for metal ions, proves that for all M(PA) complexes a larger stability is observed than is expected for a sole phosphonate coordination of the metal ion. This increased stability is attributed to the formation of five-membered chelates involving the ether oxygen present in the aliphatic residue (-CH2-O-CH2-PO3(2-)) of the ligands. The formation degrees of these chelates were calculated; they vary between about 13% for Ca(8,8aPMEA) and 71% for Cu(8,8aPMEA). The adenine residue has no influence on complex stability except in the Cu(9,8aPMEA) and Zn(9,8aPMEA) systems, where an additional stability increase attributable to the adenine residue is observed and equilibria between four different isomers exist. This means (1) an open isomer with a sole phosphonate coordination, M(PA)op, where PA(2-)=9,8aPMEA2-, (2) an isomer with a five-membered chelate involving the ether oxygen, M(PA)cl/O, (3) an isomer which contains five- and seven-membered chelates formed by coordination of the phosphonate group, the ether oxygen and the N3 site of the adenine residue, M(PA)cl/O/N3, and finally (4) a macrochelated isomer involving N7, M(PA)cl/N7. For Cu(9,8aPMEA) the formation degrees are 15, 30, 48 and 7% for Cu(PA)op, Cu(PA)cl/O, Cu(PA)cl/O/N3 and Cu(PA)cl/N7, respectively; this proves that the macrochelate involving N7 is a minority species. The situation for the Cu(PMEA) system, where PMEA2- represents the parent compound, i.e. the dianion of 9-[2-(phosphonomethoxy)ethyl]adenine, is quite similar. The relationship between the antiviral activity of acyclic nucleoside phosphonates and the structures of the various complexes is discussed and an explanation is offered why 9,8aPMEA is biologically active but 8,8aPMEA is not.     

Linkage isomerism in the metal complex hexa(thiocyanato)rhenate(IV): Synthesis and crystal structure of (NBu4)2[Re(NCS)6] and [Zn(NO3)(Me2phen)2]2[Re(NCS)5(SCN)]

The linkage isomers [Re(NCS)₆]^2? and [Re(NCS)₅(SCN)]^2? are obtained by the reaction of [ReBr₆]^2? with NCS^? in dimethylformamide. Some differences in the chemical behavior allowed their separation and structural characterization in the form of (NBu₄)₂[Re(NCS)₆] (1) and [Zn(NO₃)(Me₂phen)₂]₂[Re(NCS)₅(SCN)] (2), respectively (Bu = n-C₄H₉ and Me₂phen = 2,9-dimethyl-1,10-phenanthroline).     

The structure of a dinuclear silver(I) complex: Ag2[S2C2(CN)2] [P(C6H5)3]4

The crystal structure of the dimeric Ag maleonitriledithiolate complex, Ag₂[S₂C₂(CN)₂] [P(C₆- H₅)₃]₄ (1), has been performed. Complex 1 crystallizes in the space group P2₁/c with a = 12.2898(77), b = 23.8325(91), c = 23.1790(118) Å, β = 101.315(43)° and Z = 4. Refinement using 3253 reflections with F_o²>3σ(F_o²) yielded R = 0.0662, R_w= 0.0669. The most interesting aspect of the structure is the strong bridging interaction of the chelating maleonitriledithiolate ligand with the second Ag center, where a Ag-S distance of 2.478 Å is observed. The residual bonding capability of the sulfur atoms in the chelating anion [Ag(S₂C₂(CN)₂)(PPh₃)₂]⁻ for [Ag(PPh₃)₂]⁺ is demonstrated.     

Molecular structure of cyclo[-(D-Val-L-Hyi-L-Val-D-Hyi)2-] revealed by x-ray analysis.

The crystal structure of a synthetic analogue of valinomycin, cyclo[-(D-Val-L-Hyi-L-Val-D-Hyi)2-] (octa-meso-valinomycin) (I) (C40H68N4O12.1.5.C4H8O2, M(r) = 937.01 + 88.10), has been determined. Crystals grown from dioxane are monoclinic, space group P2(1)/a, with cell parameters a = 21.487 (8), b = 16.836 (5), c = 16.089 (4) A, beta = 111.70 (4), and Z = 4. The atomic coordinates for nonhydrogen atoms were refined in the anisotropic thermal motion approximation. H atom positions were included in the structure factor calculations at their geometrically expected positions. Values of the standard and weighted R factors after refinement are 0.11 and 0.13, respectively. The conformation of the depsipeptide crystallized from dioxane is different from that crystallized from chloroform (II). The molecule adopts a rectangular shape with two type IV beta-turns containing a hydrogen bond and possesses pseudorotational symmetry. The side chains are located on the molecular periphery. The orientation of the carbonyl groups of the molecule is not conducive for efficient metal-ion coordination and in the observed conformation cannot behave as an ionophore. In the crystal the molecules form infinite chains parallel to the c axis, and are stabilized by two intermolecular hydrogen bonds that are shorter and have better geometry than the intramolecular hydrogen bonds. A phi/psi plot for dodecadepsipeptides with a (DLLD)3 sequence has well-defined areas for Val and Hyi residues only in cases when the crystals have been grown from nonpolar or medium-polar solvents. The phi/psi plot for octadepsipeptides crystallized from chloroform (II) shows this behavior also.(ABSTRACT TRUNCATED AT 250 WORDS)     

Crystal structure of DsbDgamma reveals the mechanism of redox potential shift and substrate specificity(1)

The Escherichia coli transmembrane protein DsbD transfers electrons from the cytoplasm to the periplasm through a cascade of thiol-disulfide exchange reactions. In this process, the C-terminal periplasmic domain of DsbD (DsbDgamma) shuttles the reducing potential from the membrane domain (DsbDbeta) to the N-terminal periplasmic domain (DsbDalpha). The crystal structure of DsbDgamma determined at 1.9 A resolution reveals that the domain has a thioredoxin fold with an extended N-terminal stretch. In comparison to thioredoxin, the DsbDgamma structure exhibits the stabilized active site conformation and the extended active site alpha2 helix that explain the domain's substrate specificity and the redox potential shift, respectively. The hypothetical model of the DsbDgamma:DsbDalpha complex based on the DsbDgamma structure and previous structural studies indicates that the conserved hydrophobic residue in the C-X-X-C motif of DsbDgamma may be important in the specific recognition of DsbDalpha.     

Crystal structure of Salmonella typhimurium 2-methylisocitrate lyase (PrpB) and its complex with pyruvate and Mg(2+)

Propionate metabolism in Salmonella typhimurium occurs via 2-methylcitric acid cycle. The last step of this cycle, the cleavage of 2-methylisocitrate to succinate and pyruvate, is catalysed by 2-methylisocitrate lyase (PrpB). Here we report the X-ray crystal structure of the native and the pyruvate/Mg(2+) bound PrpB from S. typhimurium, determined at 2.1 and 2.3A, respectively. The structure closely resembles that of the Escherichia coli enzyme. Unlike the E. coli PrpB, Mg(2+) could not be located in the native Salmonella PrpB. Only in pyruvate bound PrpB structure, Mg(2+) was found coordinated with pyruvate. Binding of pyruvate to PrpB seems to induce movement of the Mg(2+) by 2.5A from its position found in E. coli native PrpB. In both the native enzyme and pyruvate/Mg(2+) bound forms, the active site loop is completely disordered. Examination of the pocket in which pyruvate and glyoxalate bind to 2-methylisocitrate lyase and isocitrate lyase, respectively, reveals plausible rationale for different substrate specificities of these two enzymes. Structural similarities in substrate and metal atom binding site as well as presence of similar residues in the active site suggest possible similarities in the reaction mechanism.     

The interaction of [Ru(NH3)5Cl]2+ and [Ru(NH3)6]3+ ions with DNA

The interaction of [Ru(NH3)5Cl]2+ and [Ru(NH3)6]3+ complex ions with calf thymus DNA has been studied at various r values (r = [Mn+]/[DNA-P]). Electronic spectra of metal-DNA solutions have been recorded and compared to the spectra of metal, as well as of DNA, solutions. Melting curves have been taken for the determination of DNA melting temperature (Tm) in the presence of the above complex ions. The results showed a biphasic melting of the DNA strands for relatively high r values. The Tm for the first phase increased with increasing r values, indicating metal ion interaction with the phosphate moieties of the DNA. The appearance of a second-phase melting, in connection with electronic spectra, pH values, and conductivity measurements of metal ion solutions, is indicative of the initial complexes' transformation to [Ru(NH3)5OH]2+, which binds preferentially to double-stranded rather than single-stranded DNA, thus leading to a second melting curve at a higher temperature than the first one.