CD Changes Research Articles

Involvement of carboxyl and phenoxyl groups in Cajanus cajan lectin-sugar interaction: interpretation of titration curves.

Hydrogen ion titration of an affinity-purified mannose/glucose-specific lectin from Cajanus cajan seeds at 30 degrees C and ionic strength of 0.15 was reported earlier by our lab (Salahuddin and Khan, 1998). Further work has been carried out on the systematic study of H+ titration of Cajanus cajan lectin at three different temperatures, i.e., 10 degrees, 20 degrees, and 30 degrees C at ionic strength of 0.15. In all, 88 protons were dissociated reversibly in pH range 2-12. Different ionizable groups present in lectin were also determined with respect to their heat of ionization. Heat of ionization, deltaH, for 43 carboxyl groups was 1.92 Kcal/mol, and for 7 phenoxyl groups, 6.84 Kcal/mol. Spectrophotometric titration was performed in the presence and absence of 0.1 M methyl alpha-D-mannopyranoside to characterize the role of phenoxyl groups present at the active site. One phenoxyl group was found to be involved in carbohydrate binding. The presence of two carboxyl groups per subunit at the active site was confirmed by H+ titration in the presence and absence of 0.1 M methyl alpha-D-mannopyranoside. Hemagglutinating and IgM precipitating activity was checked for the chemically modified carboxyl and phenoxyl group preparation. Loss of activity further confirmed the participation of carboxyl as well as phenoxyl groups at the active site. It is interesting to note the near-UV CD changes in the aromatic environment/conformation of the lectin in the presence of specific sugar, further indicating the possible participation of phenoxyl groups in the carbohydrate-binding pocket.

Pigment organization and their interactions in reaction centers of photosystem II: optical spectroscopy at 6 K of reaction centers with modified pheophytin composition.

Photosystem II reaction centers (RC) with selectively exchanged pheophytin (Pheo) molecules as described in [Germano, M., Shkuropatov, A. Ya., Permentier, H., Khatypov, R. A., Shuvalov, V. A., Hoff, A. J., and van Gorkom, H. J. (2000) Photosynth. Res. 64, 189-198] were studied by low-temperature absorption, linear and circular dichroism, and triplet-minus-singlet absorption-difference spectroscopy. The ratio of extinction coefficients epsilon(Pheo)/epsilon(Chl) for Q(Y) absorption in the RC is approximately 0.40 at 6 K and approximately 0.45 at room temperature. The presence of 2 beta-carotenes, one parallel and one perpendicular to the membrane plane, is confirmed. Absorption at 670 nm is due to the perpendicular Q(Y) transitions of the two peripheral chlorophylls (Chl) and not to either Pheo. The "core" pigments, two Pheo and four Chl absorb in the 676-685 nm range. Delocalized excited states as predicted by the "multimer model" are seen in the active branch. The inactive Pheo and the nearby Chl, however, mainly contribute localized transitions at 676 and 680 nm, respectively, although large CD changes indicate that exciton interactions are present on both branches. Replacement of the active Pheo prevents triplet formation, causes an LD increase at 676 and 681 nm, a blue-shift of 680 nm absorbance, and a bleach of the 685 nm exciton band. The triplet state is mainly localized on the Chl corresponding to B(A) in purple bacteria. Both Pheo Q(Y) transitions are oriented out of the membrane plane. Their Q(X) transitions are parallel to that plane, so that the Pheos in PSII are structurally similar to their homologues in purple bacteria.

Mechanistic Studies on the CD Degradation of 193nm Resists during SEM Inspection.

Pattern deformation during SEM inspection has been investigated for different types of 193nm positive resists. The mechanism of the CD changes is discussed based on the slimming rate analyses as well as on the e-beam penetration depth. As a general tendency, the methacrylate resists exhibit faster line width reduction than the cycloolefin-maleic anhydride (COMA) systems; however, other resist components as well as CD SEM settings play an important role. Based on the exposure time vs. CD loss, the line width slimming (LWS) is found to proceed in three steps, which are assigned as: (1) chemical change of outer resist layer, (2) evaporation of volatiles and (3) bulk chain scission or deprotection. Countermeasures for CD degradation are proposed from both the formulation and process sides. A calculation of e-beam penetration depth suggests that deprotection, chain scission and other reactions occur in the first 20-40nm, and these reaction rates combined with thermal effects determine LWS. The CD SEM measurement method has been improved to minimize e-beam exposure and to spread out the thermal load over a larger period of time. An optimized formulation exhibits less than 0.2% LWS per measurement with the improved CD measurement program.

Evaluation of Effective Image Blurring Factors in the Synchrotron Proximity X-Ray Lithography

The total effective blur, σTE, in synchrotron proximity X-ray lithography (PXRL) was evaluated to obtain a simple effective dose image. σTE characterizes the total amount of blur attributable to the source angular distribution, the exposure system, the resist material and the process. In general, the final image on the resist becomes more blurry than the aerial image due to the absorption of photons by resists and the following additional resist process such as post exposure bake (PEB). σTE2 can be understood by the square sum of many kinds of blur sources, and can be decomposed into a gap-dependent factor, seff, and a gap-independent factor, σconst. These characteristics of σTE can be evaluated using the rate of the CD changes within a gap range. We demonstrated the procedure for evaluating σTE, seff and σconst, for a specific resist material and process, along with the exposure system such as a synchrotron source and a stepper.

Comparison of PEC approaches for SCALPEL

SCALPEL is a projection electron lithography technology with resolution limited by aberrations and electron-electron interactions, not diffraction. The main CD control issue is adequate dose latitude, for image blur approaching the feature size in a high throughput scenario. This includes the effect of the unwanted electron back-scatter (EBS) proximity dose. Due to operation at 100KeV, the back-scatter range is large (∼ 25 microns) and resulting CD changes depend only on the density surroundings of a critical feature. We compare mask pattern data biasing (adjustment of binary mask object sizes) with our proposed GHOST scheme; both available as methods of proximity effect correction (PEC). We demonstrate data bias for a gate-level test pattern with realistic features and density, using a very simple model. These CD control results are confirmed with CAPROX™, a complete device-pattern PEC calculation tool from Sigma-C.

Interaction of DNA with [Cr(Schiff base)(H 2O) 2]ClO 4

The binding of Schiff base complexes of chromium(III) of the type [Cr(salen)(H 2O) 2] + and [Cr(salprn)(H 2O) 2] +, where salen denotes 1,2-bis(salicylideneamino)ethane and salprn denotes 1,3-bis(salicylideneamino)propane to calf thymus DNA has been investigated by absorption, emission, circular dichroism, melting temperature and viscosity measurements. These chromium(III) complexes showed absorption hyperchromicity accompanied by red shift in charge transfer band, fluorescence enhancement, increase in melting temperature, some structural changes in CD spectra and changes in specific viscosity when bound to calf thymus DNA. The binding constant K b has been determined from absorption measurements for both the complexes and found to be (2.5±0.4)×10 3 M −1 for [Cr(salen)(H 2O) 2] + and (1.7±0.3)×10 4 M −1 for [Cr(salprn)(H 2O) 2] +. From the binding stoichiometry of DNA-[Cr(salprn)(H 2O) 2] +, the number of binding site size has been determined and found to be ten base pairs per bound complex molecule. The chromium(III) complexes also bring about single strand cleavage in plasmid DNA. The experimental results show that the chromium(III) complexes bind to DNA by non-intercalative mode. Major groove binding is the preferred mode of interaction for these Schiff base complexes of chromium(III).

Stability, refolding and Ca2+ binding of pullulanase from the hyperthermophilic archaeon Pyrococcus woesei.

The unfolding and refolding of the extremely heat-stable pullulanase from Pyrococcus woesei has been investigated using guanidinium chloride as denaturant. The monomeric enzyme (90 kDa) was found to be very resistant to chemical denaturation and the transition midpoint for guanidinium chloride-induced unfolding was determined to be 4.86 +/- 0.29 M for intrinsic fluorescence and 4.90 +/- 0.31 M for far-UV CD changes. The unfolding process was reversible. Reactivation of the completely denatured enzyme (in 7.8 M guanidinium chloride) was obtained upon removal of the denaturant by stepwise dilution; 100% reactivation was observed when refolding was carried out via a guanidinium chloride concentration of 4 M in the first dilution step. Particular attention has been paid to the role of Ca2+ which activates and stabilizes this archaeal pullulanase against thermal inactivation. The enzyme binds two Ca2+ ions with a Kd of 0.080 +/- 0.010 microM and a Hill coefficient H of 1.00 +/- 0.10. This cation enhances significantly the stability of the pullulanase against guanidinium chloride-induced unfolding and the DeltaGH2OD increased from 6.83 +/- 0.43 to 8.42 +/- 0.55 kcal.mol-1. The refolding of the pullulanase, on the other hand, was not affected by Ca2+.

The effect of long-term phosphatic fertiliser applications on the amounts and forms of cadmium in soils under pasture in New Zealand

This paper presents the results of an investigation into the rate of Cd accumulation and changes in forms of Cd in a soil that has been subjected to long-term superphosphate fertiliser application. Results indicate that there had been a significant accumulation of Cd in the soil during the past 44 years. On the high fertiliser treatment (376 kg superphosphate ha-1 yr-1), Cd was estimated to have accumulated at a rate of 7.8 g ha-1 yr-1. During the course of the trial, there was an increase in the proportion of Cd associated with exchangeable and soil organic matter fractions on the fertilised plots, which was related to a corresponding increase in soil organic carbon levels. Results also indicate that although there was a large proportion of added Cd associated with the organic fraction, the concentration of applied Cd occurring in the residual fraction was also substantial (i.e >25% of added Cd). In addition, an investigation into the effects of residence time of Cd in the soil indicated that there was a redistribution of Cd into less soluble forms with time (i.e. residual Cd), along with a decrease in total soil Cd concentrations. There was also evidence of movement of Cd down the soil profile in this irrigated soil. The implications of these results for Cd phytoavailability are discussed.

Effect of temperature on DNA secondary structure in the absence and presence of 0.5 M tetramethylammonium chloride.

Changes in the average secondary structures of three different linear DNAs over the premelting region from 5 to 60 degrees C were investigated by measuring their CD spectra and also their torsion elastic constants (<alpha>) by time-resolved fluorescence polarization anisotropy. For one of these DNAs, the Haell fragment of pBR322, the apparent diffusion coefficients [Dapp(k)] at small and large scattering vectors (k) were also measured by dynamic light scattering. With increasing temperature, all three DNAs exhibited typical premelting changes in their CD spectra, and these were accompanied by 1.4- to 1.7-fold decreases in <alpha>. Also for the 1876 base pair fragment, Dapp(k) at large scattering vectors, which is sensitive to the dynamic bending rigidity, decreased by 17%, even though there was no change at small scattering vectors, where Dapp(k) = D0 is the translational diffusion coefficient of the center-of-mass. These observations demonstrate conclusively that the premelting CD changes of these DNAs are associated with a significant change in average secondary structure and mechanical properties, though not in persistence length. In the presence of 0.5 M tetramethylammonium chloride (TMA-Cl) the premelting change in CD is largely suppressed, and the corresponding changes in <alpha> and Dapp(k) at large scattering vectors are substantially diminished. These observations suggest that TMA-Cl, which binds preferentially to A.T-rich regions and stabilizes those regions (relative to G.C-rich regions) against melting, effectively stabilizes the prevailing low-temperature secondary structure sufficiently that the DNA is effectively trapped in that state over the temperature range observed.

Binding of 2-azaanthraquinone derivatives to DNA and their interference with the activity of DNA topoisomerases in vitro.

We have investigated the binding ability to DNA of compounds belonging to the 2-azaanthraquinone-type structure and have examined the effect on the activity of DNA gyrase as well as on mammalian topoisomerases in vitro. Using different biophysical techniques it was found that one of these ligands, 9-((2-dimethylamino)ethyl)amino)-6-hydroxy-7-methoxy-5, 10-dihydroxybenzo[g]isoquinoline-5,10-dione (TPL-I), is an intercalating DNA binding agent, whereas the parent compound tolypocladin (TPL) and a derivative (TPL-II) showed almost no similar affinity to DNA. CD measurements demonstrated a significant and selective binding tendency of TPL-I to alternating purine/pyrimidine sequences with some preference for poly(dA-dT). poly(dA-dT). Tm values were increased of the ligand complex with the alternating AT-containing duplex polymer. The binding to various DNAs was characterized by CD and visible absorption spectral changes. From the latter, different binding constants of 6.2 x 10(5) and 1.5 x 10(5) M-1 were obtained for poly(dA-dT).poly(dA-dT) and poly(dA). poly(dT), respectively. Sedimentation measurements with supercoiled pBR322 plasmid DNA clearly indicated an intercalative binding mechanism associated with an unwinding angle of about 18 degrees. These results suggest that the intercalative binding of TPL-I is promoted by the 2-(dimethylamino)ethylamino group substituted on carbon 9 of the anthraquinone system. The cytotoxic compound TPL-I, but not TPL or TPL-II, effectively inhibited the DNA supercoiling reaction of DNA gyrase and the activity of mammalian topoisomerases I and II as measured by the relaxation assay. TPL-I affects the cleavage reaction of topoisomerases on a single site located in alternating purine-pyrimidine sequence regions. The inhibitory potency of TPL-I can be ascribed to a blocking of cleavage sites on the DNA substrate, which correlates with the sequence preference of the ligand.

Interaction of oxytocin with Ca2+: I. CD and fluorescence spectral characterization and comparison with vasopressin

Extracellular Ca2+ is required for the action of oxytocin and both the hormone and its receptor have binding sites for divalent metal cations. To characterize the cation-bound form of oxytocin, we monitored the binding of Ca2+ and Mg2+ to oxytocin as well as peptides representing its ring and tail regions in trifluoroethanol, a lipid-mimetic solvent, using CD and fluorescence spectroscopy. Binding Ca2+ (Kd ˜ 50 μM) caused drastic CD and fluorescence changes leading to a helical conformation. Mg2+ caused CD changes smaller than and opposite to Ca2+. However, the helical structure was enhanced when both Ca2+ and Mg2+ were present together. CD changes in the tail peptide of oxytocin showed its ability to bind Ca2+ and Mg2+ whereas the vasopressin tail peptide did not bind either cation. CD spectral changes on Ca2+ and Mg2+ binding to tocinoic acid (the ring moiety of oxytocin) were much smaller than those of oxytocin. These data suggest that the tail segment of oxytocin potentiates Ca2+ binding by the ring. While vasopressin displayed a CD spectrum similar to that of oxytocin, CD spectra of its cation-bound forms were markedly different from those of oxytocin; the Ca2+-induced CD changes in vasopressin were very much smaller and of opposite sign, and Mg2+-induced ones significantly larger than in oxytocin. Taken together, our observations bring out the structural differences between oxytocin and vasopressin in the context of their interaction with Ca2+ and Mg2+. This may be relevant to understanding the differences in the bioactive conformations and receptor interactions of the two hormones. © 1997 John Wiley & Sons, Inc. Biopoly 40: 433–443, 1996

Interaction of oxytocin with Ca2: I. CD and fluorescence spectral characterization and comparison with vasopressin

Extracellular Ca2+ is required for the action of oxytocin and both the hormone and its receptor have binding sites for divalent metal cations. To characterize the cation-bound form of oxytocin, we monitored the binding of Ca2+ and Mg2+ to oxytocin as well as peptides representing its ring and tail regions in trifluoroethanol, a lipid-mimetic solvent, using CD and fluorescence spectroscopy. Binding Ca2+ (Kd ˜ 50 μM) caused drastic CD and fluorescence changes leading to a helical conformation. Mg2+ caused CD changes smaller than and opposite to Ca2+. However, the helical structure was enhanced when both Ca2+ and Mg2+ were present together. CD changes in the tail peptide of oxytocin showed its ability to bind Ca2+ and Mg2+ whereas the vasopressin tail peptide did not bind either cation. CD spectral changes on Ca2+ and Mg2+ binding to tocinoic acid (the ring moiety of oxytocin) were much smaller than those of oxytocin. These data suggest that the tail segment of oxytocin potentiates Ca2+ binding by the ring. While vasopressin displayed a CD spectrum similar to that of oxytocin, CD spectra of its cation-bound forms were markedly different from those of oxytocin; the Ca2+-induced CD changes in vasopressin were very much smaller and of opposite sign, and Mg2+-induced ones significantly larger than in oxytocin. Taken together, our observations bring out the structural differences between oxytocin and vasopressin in the context of their interaction with Ca2+ and Mg2+. This may be relevant to understanding the differences in the bioactive conformations and receptor interactions of the two hormones. © 1997 John Wiley & Sons, Inc. Biopoly 40: 433–443, 1996

Circular dichroism spectroscopy of three tyrosine-to-phenylalanine substitutions of fd gene 5 protein.

Circular dichroism spectroscopy was used to study mutants of phage fd gene 5 protein (Y26F, Y34F, and Y41F) in which three of the five tyrosines, Tyr-26, Tyr-34, and Tyr-41, were individually substituted with phenylalanine. The tyrosine 229 nm CD bands of the wild type, Y26F, and Y41F gene 5 proteins decreased in magnitude during complex formation with either fd ssDNA or poly[d(A)]. However, the Y34F gene 5 protein showed no decrease in the 229 nm band during titrations of these nucleic acids. This suggested that Tyr-34 of the wild type gene 5 protein dominated the 229 nm CD changes upon binding to single-stranded DNA. Titrations of poly[d(A)] or fd ssDNA with wild type, Y26F, Y34F, or Y41F gene 5 proteins resulted in perturbations of the nucleic acid near-UV CD bands, specific for the particular nucleic acid, but similar for all four proteins (in 2 mM sodium phosphate buffer, pH 7.0). For both nucleic acids, the addition of protein beyond a certain [protein monomer]/[nucleotide] ratio (0.25 for poly[d(A)] or 0.33 for fd ssDNA) resulted in a partial reversal of the CD change of the nucleic acid. These data are interpreted to mean that, in addition to the two well-known n = 4 and n = 3 stoichiometric modes of binding, there is a third mode of binding in which the nucleic acid is in limited contact with the protein. As shown by salt dissociation studies of complexes with poly[d(A)], the binding affinities, K omega, of the proteins were in the order: wild type > Y26F >> Y34F > or = Y41F (for the n = 4 binding mode in 0.1-0.2 M NaCl). Our data indicate that Tyr-34 plays a more important role in forming a complex with ssDNA than is apparent in current models of the g5p.ssDNA complex. We suggest that the hydroxyl moieties of Tyr-34 and Tyr-41 are both somehow involved in stabilizing the interface of bound protein dimers.

The Molybdenum Centers of Xanthine Oxidase and Xanthine Dehydrogenase: DETERMINATION OF THE SPECTRAL CHANGE ASSOCIATED WITH REDUCTION FROM THE Mo(VI) TO THE Mo(IV) STATE

The UV-visible absorbance change associated with reduction of the molybdenum centers of xanthine oxidase and xanthine dehydrogenase has been determined using a double-difference technique. At pH 8.5, the Mo(VI) minus Mo(IV) difference spectrum seen with xanthine oxidase exhibits a positive feature at 420 nm, having an extinction change of approximately 3,000 M-1 cm-1 as well as evidence for a negative feature below 340 nm. In xanthine oxidase this change is found to exhibit a marked pH dependence, implicating protonation/deprotonation events associated with changes in the molybdenum oxidation state. Application of the double-difference protocol to the respective circular dichroism spectra of xanthine oxidase and xanthine dehydrogenase reveals appreciable CD changes at 420 and 580 nm associated with the reduction of the molybdenum center. The present results demonstrate a direct spectroscopic handle on the molybdenum centers of both xanthine oxidase and xanthine dehydrogenase.

Extraction studies of Zn(II), Cu(II) and Cd(II) with impregnated and Levextrel resins containing di(2-ethylhexyl) phosphoric acid (Lewatit 1026 Oc)

Impregnated resins are an alternative approach to the usual techniques for the recovery of metal ions, such as solvent extraction and ion exchange resins. This paper presents the results of the extraction of Zn(II), Cu(II) and Cd(II) from nitrate and chloride solutions at 0.1 M ionic strength and 25°C by the Levextrel impregnated resin Lewatit 1026 Oc containing di(2-ethylhexyl)-phosphoric acid (DEHPA). The distribution coefficients were determined as a function of both pH and ionic medium and the data were analyzed graphically, using the slope analysis method, and numerically using the program letagrop-distr. Analysis of the results shows that the extraction of these metal ions can be explained assuming the formation of metal complexes in the resin phase have a general composition ML 2(HL) q where q takes different values depending on the metal. An extraction reaction is proposed and the extraction constants of the species are given. The adsorption of DEHPA molecules on the styrene/divinyl benzene macroporous support is due to weak extractant-support interactions, as indicated by FTIR spectroscopy. Furthermore, it is also shown that the ability of the extractant towards Zn(II), Cu(II) and Cd(II) changes in the solid support, in comparison with organic solvents. Finally, a comparison is made between the extraction of Zn(II), Cu(II) and Cd(II) with Lewatit 1026 Oc resins and impregnated resins prepared by direct adsorption of DEHPA into Amberlite XAD2 (XAD2-DEHPA resins).

Unfolding of 4-aminobutyrate aminotransferase equilibrium and kinetic studies

The unfolding of pig liver 4-aminobutyrate aminotransferase by urea has been investigated at equilibrium. The overall process was reversible as judged from the recovery of catalytic activity after dilution of urea-treated samples. Unfolding of the enzyme was monitored by circular dichroism and fluorescence spectroscopy. The steepness of the fluorescence and CD changes between 2 and 8 M urea, and the lack of any discernible plateau suggests that unfolding of the protein is a cooperative process. The unfolding of 4-aminobutyrate aminotransferase as a function of urea concentration was monitored by fluorescence measurements of the tryptophanyl residues. The kinetic results indicate that the aminotransferase unfolds in a single kinetic phase. Unfolded 4-aminobutyrate aminotransferase recovers immediately its catalytic activity upon dilution with buffers of neutral pH. Based on equilibrium and kinetic results, a two- state model for the unfolding of the aminotransferase is proposed.

Precipitation and Post-Depositional Mobility of Cadmium in Reducing Sediments: Implications for the Cd Oceanic Budget

Reconstructions of past nutrient distribution and deep ocean circulation patterns are typically inferred from the 513C and Cd composition of benthic foraminifera from deep sea sediments; the modern oceanic distribution of 5 t3C and Cd reflect the biological cycling of nutrients (e.g. PO4) as modified by deep ocean circulation. This correlation between Cd and PO4 is the basis for using Cd, as recorded in foraminiferal shells, for reconstructing changes in deepwater circulation patterns (Boyle, 1988). Because of the relatively long residence time of Cd and PO4 (100,000-200,000 years) glacial-interglacial changes in Cd can be interpreted as reflecting variations in nutrients distribution. However, Cd and PO4 are controlled by different diagenetic processes. The strong tendency of Cd to form highly insoluble sulphide phases in the presence of HES explains its substantial enrichment in sulphide-bearing sediments as already was noted by Goldschmidt (1958). In recent studies we have investigated the possibility that temporal changes in the areal extent of reducing sediments may affect the Cd oceanic budget thus leading to de-coupling of Cd from the nutrient cycle (Rosenthal et al., 1993). The current work focuses on the precipitation of authigenic Cd in suboxic sediments and its mobility in response to post-depositional reoxidation of reducing sediments. The implications of these processes for the global Cd budget and for paleoceanographic interpretation are discussed below.

Effect of Hg(II) on d(GCGCATATGCGC) 2 conformation: UV absorption and circular dichroism studies

Exposing the palindromic dodecamer d(GCGCATATGCGC) 2, dissolved in the neutral-salt solvent 0.1 M (≡ low salt) or 3.0 M (≡ high salt) NaClO 4, to increasing amounts of Hg(ClO 4) 2 (≡ Hg(II)) produces major changes in its chirality in the low-salt medium but not high-salt. Let r ≡ [Hg(ClO 4) 2] added/[DNA-P]. At low salt, the conformational changes are noted as follows; in absence in Hg(II), the near-conservative CD spectrum of the dodecamer is that of a B-type DNA. In the presence of Hg(II), the long-wavelength positive Cotton band (central location near 280 nm) undergoes inversion at r > 0.2 and reaches maximum negative extension at r = 0.5–0.6. A new minor positive CD peak appears around 315 nm. The long-wavelength negative Cotton band (central location near 250 nm) experiences a slight red-shift by about 10 nm but remains otherwise rather unaffected by Hg(II). Major CD changes are also noted in the short-wavelength region near 220 nm. The inverted CD spectrum is in general more complex in structure than the spectrum of the untreated oligomer. By contrast, the CD changes seen in the high-salt medium are not as complex, and the CD scripture of a right-handed B-type helix is retained. It is stressed that all CD changes are fully reversible: both the low-salt and high-salt CD spectra of the untreated dodecamer are regained subsequent to the removal of Hg(II) with the help of a complexing agent such as sodium cyanide. The (ordinary) UV absorption spectra of the dodecamer are altered by Hg(II) in less spectacular fashion although differences in progression in the two-salt media are noted. We interpret the CD changes as signaling (right-handed) B ↔ (left-handed) non-B transitions of the oligodeoxyribonucleotide at low-salt levels but not at high-ionic strength. The presence of isodichroic (isosbestic) points in the various spectra shows that the Hg(II)-induced conformational transitions are governed by well-defined equilibria. Whether the CD changes represent B ↔ Z transitions is unclear at present.

The cholesterol-side-chain-cleaving cytochrome P450 spin-state equilibrium. 2. Conformational analysis.

We have confirmed and characterised structurally the enzyme conformational changes deduced from the preceding thermodynamic analysis of the adrenal mitochondrial cholesterol-side-chain-cleaving cytochrome P450 spin-state equilibrium. The spin-transition kinetics following rapid pH jumps were multiphasic in aqueous buffer and biphasic in the presence of 35% ethylene glycol. The activation energy between -2 degrees C and 30 degrees C of both phases was exceptionally high (Ea = 147 kJ.mol-1), suggesting the involvement of large-scale conformational changes. The pH and temperature effects on the CD spectrum show that the enzyme is in equilibrium between at least two conformations which are predicted by the thermodynamic model, but which are not directly correlated to the spin state. The CD changes between 260 nm and 280 nm indicate that the conformation prevailing at high temperatures is characterised by a decreased polarity of the tyrosine environments; the changes between 200 nm and 250 nm suggest furthermore a 4% decreased protein helical content.

Calcium deficiency enhances cadmium accumulation in the central nervous system

Weanling male rats were administered 1 of 4 diets for 40 days: control (CONT), low Ca (LOCA), control plus Cd (CONT+Cd) or low Ca plus Cd (LOCA+Cd). After 40 days, Cd was analyzed in 7 brain regions, spinal cord, serum, liver, kidney; muscle and femur by atomic absorption spectrophotometry with Zeeman background correction. No significant difference in Cd between CONT and LOCA was found except in femur, where it was increased. In CONT+Cd rats, peripheral tissues showed an increase in Cd of 30–71 fold above CONT rats. Brain regions exhibited a more modest 7–10 fold change, and serum Cd was 8.5 times above control. LOCA+Cd rats showed a 25-fold increase of Cd above LOCA in serum, 25–100 fold in peripheral tissues, and a 14–20 times increase in brain. These findings show that brain Cd is increased during Ca deficiency, but that central nervous system Cd changes less than peripheral tissue Cd. This increase in brain Cd could after brain function.