A conformational analysis of mono and dialkyl ethers of 2,2′-dihydroxy-1,1′-binaphthalene by circular dichroism spectroscopy and cholesteric induction in nematic liquid crystals

The coupling of the analysis of the absorption and circular dichroism (CD) spectra with that of the cholesteric mesophases induced in nematic liquid crystals indicated some interesting conformational features of bridged and nonbridged mono- and dialkylethers of optically active 2,2′-dihydroxy-1,1′-binaphthalene. Bridged derivatives are characterized by relatively small dihedral angles. Simple monoalkyl ethers are characterized by larger dihedral angles but they all assume an s-cis conformation, owing to the existence of intramolecular hydrogen bonds. Nonbridged dialkylethers prefer even larger dihedral angles and, depending on the bulkiness of the alkyl groups, the s-trans conformation can be found. Interestingly, the conformation of dialkylethers is strongly dependent on the structure of the liquid crystal solvent, because the intramolecular hydrogen bond is not possible there. © 1995 Wiley-Liss, Inc.     

Conformational analysis of bioactive peptides in reverse micelles as mimics of cell membrane environments

Summary Conformational preferences of secretin as a model peptide have been analyzed by CD and IR spectroscopy in reverse micelles of AOT/isooctane/water and compared to those in aqueous TFE, in SDS micelles and in DMPG vesicles. Among the systems examined, reverse micelles and phospholipid vesicles displayed almost identical conformational equilibria. Very high lipid-to-peptide ratios can be obtained in reverse micelles with full retention of optical transparency, even at millimolar peptide concentrations, thus indicating this system to be an interesting mimic of cell membrane environments for spectroscopic analysis of bioactive peptide conformations.Abbreviations TFE trifluoroethanol - SDS sodium dodecyl sulfate - DMPG dimyristoylphosphatidylglycerol - AOT bis(2-ethylhexyl)sulfosuccinate - CMC critical micellar concentration - VIP vasoactive intestinal peptide     

Two steps in the transition between the native and acid states of bovine α-lactalbumin detected by circular polarization of luminescence: Evidence for a premolten globule state?

A few studies indirectly support the existence of an intermediate in the transition of Ca(2+)-saturated bovine alpha-lactalbumin (alpha-LA) from the native (N) to the acidic (A) state, known as the molten globule state. However, direct experimental evidence for the appearance of this intermediate has not been obtained. The signal of circular polarization of luminescence (CPL) is sensitive to fine conformational transitions because of its susceptibility to changes in the environmental asymmetry of fluorescent chromophores in their excited electronic states. In the present study, CPL measurements were applied using the intrinsic tryptophan fluorescence of alpha-LA as well as the fluorescence of 8-anilino-1-naphthalenesulfonic acid (ANS) bound to alpha-LA. CPL of tryptophan and ANS was measured in the pH range of 2.5-6 in order to find direct experimental evidence for the proposed intermediate. CPL (characterized by the emission anisotropy factor, g(em)) depends on the asymmetry of the protein molecular structure in the environment of the tryptophan and the ANS chromophores in the excited electronic state. The pH dependence of both the gab, absorption anisotropy factor determined by CD, and the ANS steady state fluorescence, showed a single transition at pH 3-3.7 as already reported elsewhere. This transition was interpreted as being a result of a change of the alpha-LA tertiary structure, which resulted in a loss of asymmetry of the environment of both the tryptophan residues and the ANS hydrophobic binding sites. The pH dependence of the tryptophan and ANS g(em) showed an additional conformational transition at pH 4-5, which coincided with the pKa of Ca2+ dissociation (pKa 5), as predicted by Permyakov et al. (1981, Biochem Biophys Res Commun 100:191-197). The titration curve showed that there is a pH range between 3.7 and 4.1 in which alpha-LA exists in an intermediate state between the N- and A-state. We suggest that the intermediate is the premolten globule state characterized by a reduced Ca2+ binding to the alpha-LA, native-like tertiary structure, and reduced asymmetric fluctuation of the tertiary structure on the nanosecond time scale. This intermediate resembles the "critical activated state" theoretically deduced by Kuwajima et al. (1989, J Mol Biol 206:547-561). The present study demonstrates the power of CPL measurements for the investigation of folding/unfolding transitions in proteins.     

高原鼠兔与达乌尔鼠兔食物资源维生态位的研究

作者于１９９３年在青海省刚察县年诺索玛地区发现高原鼠兔（Ｏｃｈｏｔｏｎａ ｃｕｒｚｏｎｉａｅ）和达乌尔鼠兔（０．ｄａｕｒｉｃａ）在布哈河谷二阶同域且重叠分布之现象,并从这两种鼠兔的食物资源利用出发,研究了其生态位关系问题。在食物资源维上的生态位宽度（ＰＳ）,高原鼠兔为０．４７４４,达乌尔鼠兔为０．４９６４,即它们具有相近似的生态位宽度。这两种动物的生态位在食物资源谱维上的重叠值（ＦＴ）为０．８０９     

脂质泡中的菌紫质旋转运动的介质依赖性

用闪光诱导瞬间二向色性方法测量了蔗糖和甘油对菌紫质（ＢＲ）分子在脂质泡中的旋转扩散运动的影响。结果表明脂质囊泡本身在悬浮液中的运动并不会影响ＢＲ分子在膜中的旋转运动的测量。蔗糖和甘油对ＢＲ旋转运动的影响在相变温度以上和相变温度以下是不同的,相变温度以下的主要作用是相分离,使运动减慢,相变温度以上的作用可能是脂的分散,使运动加快。     

Orientation of photosystem-I pigments. Investigation by low-temperature linear dichroism and polarized fluorescence emission

Using absorption and fluorescence experiments at low temperature with polarized light on oriented samples, the orientation of PS-I-related pigments, both in green plants and in Chlamydomonas reinhardtii, has been investigated on isolated pigment-protein complexes and intact thylakoids. The following observations have been made. (i) The isolation procedure of PS I₁₁₀, PS I₆₅, LHC I and CP0) particles from pea and C. reinhardtii do not alter significantly the intrinsic orientation of the pigments inside the complexes; (ii) Chl b is a structural component of PS I, linked to the peripheral antenna, with an orientation with respect to the thylakoid plane different from that observed in the main light-harvesting complex (iii) PS I₆₅ (i.e., ‘core’ PS I) of pea and C. reinhardtii contains identical chromophores having the same orientation with respect to the geometrical longest axis (axes) of the complexes. (iv) LHC I and CP0 (i.e., PS I ‘peripheral antenna’) of pea and C. reinhardtii have identical oriented chromophores, except that a long-wavelength component with a high anisotropy is only present in green plants. This set of pigments, which absorbs at 705–725 nm, has the same orientation as the dipoles emitting F₇₃₅ and also as the Q_Y transition of P-700. (v) All the long-wavelength fluorescence properties of the various studied membranes are explained by these data on isolated PS I complexes: wild-type C. reinhardtii and Chl-b-less barely fluoresce from the core pigments, while a CP1 deficient mutant of C. reinhardtii and wild-type barley fluoresce from the antenna pigments.     

A thin quartz cell suitable for vacuum ultraviolet absorption and circular dichroism measurements

The design of a thin quartz cell suitable for absorption and circular dichroism measurements in the vacuum ultraviolet is described. Important features of the cell are (1) that it can be disassembled for cleaning and reproducibly reassembled with path lengths up to 0.3 mm, and (2) that strain in the windows from the compressed sample can be relieved by a sample overflow port. The latter feature allows the cell to be used for circular dichroism as well as absorption measurements.     

Dynamic structure of metallothionein

Molecular sieve chromatography of rabbit liver metallothionein at different electrolyte concentrations revealed that this protein undergoes an increase in Stokes radius from 1.50 to 1.78 nm when the ionic strength is lowered from 0.5 to 0.015 indicating a change in molecular shape and/or hydration. The variation in ionic strength also affects the far-UV circular dichroism of metallothionein reflecting a conformational transition in the protein. The effects are attributed to changes in intramolecular repulsion between the strongly negatively charged metal-thiolate clusters of the protein. It is suggested that metallothionein exists in at least two interchangeable conformational states which differ in hydrodynamic properties and whose equilibrium concentrations are determined by the electrostatic free energy of the system.     

The reaction of cysteine with ceruloplasmin copper

The kinetics of decay in absorbance at 610 nm in the reaction of cysteine with ceruloplasmin was biphasic under anaerobic conditions. Admission of oxygen to the bleached ceruloplasmin restored the blue color to about 75 % of the original value. However, under aerobic or anaerobic conditions an initial bleaching corresponded to a 25 % decrease in blue color. This change was irreversible and remained after removal of excess cysteine from the reaction mixture by dialysis. There was no correlation between transient and steady-state kinetic parameters. Circular dichroism measurements showed a characteristic reduction in the negative band at 450 nm, which is specific for type 1b copper. Isolation and further studies on cysteine-modified ceruloplasmin with a lower A₆₁₀/A₂₈₀ ratio showed < 10% reduction in enzyme activity toward p-phenylenediamine and o-dianisidine. Evidence is also presented that ceruloplasmin catalyzes the oxidation of cysteine with a one-electron reduction of oxygen and the formation of superoxide ion, which is then converted to H₂O₂ by ceruloplasmin. The effect of superoxide dismutase and catalase also confirms the presence of superoxide and H₂O₂. In sum, these data show that a permanent reduction of type 1b copper occurred when cysteine was used as a substrate. We conclude that there is a single electron transfer from cysteine directly to oxygen using one specific copper of ceruloplasmin, type 1b.     

Cellular and subcellular distribution of iron in the lamina propria of rat duodenum

Summary Cellular and subcellular distribution of iron in the lamina propria of rat duodenum was studied after a single i.p. injection of iron dextran, using electron microscopy and peroxidase cytochemistry. X-ray spectrum microanalysis was used for positive identification of iron. Ironcontaining particles (IP) were found in the cytoplasm of three cell types, viz. macrophages, pericytic reticular cells and sheathing fibrocytes. IP-containing organelles in lamina propria cells were more heterogeneous compared to absorptive cells and, in addition, some differences were noted in the subcellular distribution of IP in the 3 cell types. A common denominator in these 3 cell types was the presence of endogenous peroxidase, also shared by Kupffer cells which are known to be involved in iron storage. Peroxidase activity was absent in absorptive epithelial cells. It is hypothesized that the cells of the lamina propria, like Kupffer cells, may be the site of storage of excess iron absorbed, releasing iron upon demand and migrating into the lumen to prevent iron overload. In this fashion they may regulate the exchange of iron with the environment. The presence of peroxidase in these as well as Kupffer cells, and its absence in absorptive cells also raises the possibility that this enzyme may be related to certain aspects of iron storing process.