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ABSTRACT: A pH-responsive rolled-sheet morphology was prepared from a triskelion A(2)B-type amphiphilic polypeptide having a histidine residue as a pH-responsive unit. The dimensions of the rolled sheet were 85 nm diameter and 210 nm length with a sheet turn number of 2.0 at pH 7.4. Upon decreasing the pH from 7.4 to 5.0, the layer spacing of the rolled sheets was widened from ca. 9 to ca. 19 nm due to electrostatic repulsion caused by histidine protonation. This morphology change occurred reversibly with a pH change between 7.4 and 5.0. The molecular packing in the rolled sheets was shown to be loosened at pH 5.0 on the basis of electron diffraction measurements. The tightness of the rolled sheets was thus controlled reversibly by a pH change due to a single protonation in the amphiphilic polypeptide.
Langmuir 03/2012; 28(14):6006-12. · 4.19 Impact Factor
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ABSTRACT: In this paper, we report the combination of a near-infrared (NIR) spectroscopic method with multivariate analysis in order to develop a calibration model of the saccharification ratio of chemically pretreated Erianthus. The regression models clearly depend on the NIR spectral regions, and the information of CH and aromatic framework vibrations contributed most effectively to the alkaline dataset. From interpretations of the regression coefficient, lignin and cellulose were negatively and positively correlated with the saccharification ratio, respectively, and this result was supported by the data from wet chemical analysis. A more complex dataset was obtained from varied chemical pretreatments; here, the saccharification ratio was either small or had no linear correlation with each structural monocomponent. These results enabled the successful construction of the PLS regression model. NIR spectroscopy can be a rapid screening method for the saccharification ratio, and furthermore, can provide information of the key factors influencing the realization of more efficient enzymatic accessibility.
Applied biochemistry and biotechnology 11/2011; 166(3):711-21. · 1.94 Impact Factor
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ABSTRACT: This study reinvestigated the synthesis of cellulose in vitro with a well-known cellulose-producing bacterium, Gluconacetobacter xylinus. Alkylmaltoside detergents, which are more frequently used in recent structural biological researches, are uniquely used in this study to solubilize cellulose-synthesizing activity from the cell membrane of G. xylinus. Activity comparable to that previously reported is obtained, while the synthesized cellulose is crystallized into a non-native polymorph of cellulose (cellulose II) as well as the previous studies. In spite of this failure to recover the native activity to synthesize cellulose I microfibril in vitro, the product is a polymer with a degree of polymerization greater than 45 as determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). It was thus concluded that the established protocol can solubilize cellulose-synthesizing activity of G. xylinus with polymerizing activity.
Carbohydrate research 10/2011; 346(17):2760-8. · 2.03 Impact Factor
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ABSTRACT: Vesicles prepared from a mixture of (Sar)(25)-b-(L-Leu-Aib)(6) (SLL) and (Sar)(25)-b-(D-Leu-Aib)(6) (SDL) fused with themselves upon heating to 90 °C. The vesicles also fused with (Sar)(28)-b-(L-Leu-Aib)(8) vesicles upon heating to 90 °C. The temperature-triggered fusion was due to the phase transition of the mixed membrane of SLL and SDL at 90 °C and should be driven by the bending energy stored in the stereocomplex membrane upon taking a vesicular structure.
Langmuir 03/2011; 27(8):4300-4. · 4.19 Impact Factor
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ABSTRACT: Two types of peptide nanotubes, one is prepared from an amphiphilic peptide having a right-handed helix segment and the other from that having a left-handed helix segment, are shown to transform the morphology into a vesicle by membrane fusion due to stereo-complex formation between these helical segments.
Chemical Communications 02/2011; 47(11):3204-6. · 6.17 Impact Factor
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ABSTRACT: Amphiphilic helical peptides (Sar)(m) -b-(L-Leu-Aib)(n) (m = 22-25; n = 7, 8, 10) with a hydrophobic block as a right-handed helix were synthesized and their mixtures with (Sar)(25) -b-(D-Leu-Aib)(6) containing the hydrophobic block as a left-handed helix were examined in their molecular assembly formation. The single component (Sar)(25) -b-(D-Leu-Aib)(6) forms peptide nanotubes of 70 nm diameter and 200 nm length. The two-component mixtures of (Sar)(25) -b-(D-Leu-Aib)(6) with (Sar)(24) -b-(L-Leu-Aib)(7) , (Sar)(22) -b-(L-Leu-Aib)(8) , and (Sar)(25) -b-(L-Leu-Aib)(10) yield peptide nanotubes of varying dimensions with 200 nm diameter and 400 nm length, 70 nm diameter and several micrometer length (maximum 30 µm), and 70 nm diameter and 100-600 nm length, respectively. The right-handed and the left-handed helix were thus found to be molecularly mixed due to the stereo-complex formation and to generate nanotubes of different sizes. When the mismatch of the hydrophobic helical length between the two components was of four residues, the longest nanotube was generated. Correspondingly, the hydrophobic helical segments have to interdigitate with an anti-parallel orientation at the hydrophobic core region of the nanotube.
Journal of Peptide Science 02/2011; 17(2):94-9. · 1.80 Impact Factor
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ABSTRACT: We report a simple analytical procedure combining near-infrared (NIR) spectroscopy with multivariate analysis to detect the saccharification efficiency of pretreated rice straw. Three types of sample preparation methods were tested to develop a powerful calibration model, with the disk sample used as the standard protocol. From the spectra dataset of NaOH-treated biomass, we obtained a good calibration for the saccharification ratio and some major structural components by partial least-squares regression. Adding dataset from hot water and dilute sulfuric acid pretreatments to NaOH sample dataset, an acceptable calibration model to predict the saccharification ratio as well as the glucose, xylose, and lignin contents was generated. NIR has a great potential for rapid screening of saccharification efficiency of pretreated biomass, which would allows us to control the quality of processing toward better bioethanol production.
Applied biochemistry and biotechnology 12/2010; 164(2):194-203. · 1.94 Impact Factor
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ABSTRACT: Prokaryotic voltage-gated sodium channels (Na(V)s) form homotetramers with each subunit contributing six transmembrane α-helices (S1-S6). Helices S5 and S6 form the ion-conducting pore, and helices S1-S4 function as the voltage sensor with helix S4 thought to be the essential element for voltage-dependent activation. Although the crystal structures have provided insight into voltage-gated K channels (K(V)s), revealing a characteristic domain arrangement in which the voltage sensor domain of one subunit is close to the pore domain of an adjacent subunit in the tetramer, the structural and functional information on Na(V)s remains limited. Here, we show that the domain arrangement in NaChBac, a firstly cloned prokaryotic Na(V), is similar to that in K(V)s. Cysteine substitutions of three residues in helix S4, Q107C, T110C, and R113C, effectively induced intersubunit disulfide bond formation with a cysteine introduced in helix S5, M164C, of the adjacent subunit. In addition, substituting two acidic residues with lysine, E43K and D60K, shifted the activation of the channel to more positive membrane potentials and consistently shifted the preferentially formed disulfide bond from T110C/M164C to Q107C/M164C. Because Gln-107 is located closer to the extracellular side of helix S4 than Thr-110, this finding suggests that the functional shift in the voltage dependence of activation is related to a restriction of the position of helix S4 in the lipid bilayer. The domain arrangement and vertical mobility of helix S4 in NaChBac indicate that the structure and the mechanism of voltage-dependent activation in prokaryotic Na(V)s are similar to those in canonical K(V)s.
Journal of Biological Chemistry 12/2010; 286(9):7409-17. · 4.77 Impact Factor
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ABSTRACT: Voltage-sensor domains (VSDs) in voltage-gated ion channels are thought to regulate the probability that a channel adopts an open conformation by moving vertically in the lipid bilayer. Here we characterized the movement of the VSDs of the prokaryotic voltage-gated sodium channel, NaChBac. Substitution of residue T110, which is located on the extracellular side of the fourth transmembrane helix of the VSD, by cysteine resulted in the formation of a disulfide bond between adjacent subunits in the channel. Our results suggest that T110 residues in VSDs of adjacent subunits can come into close proximity, implying that the VSDs can move laterally in the membrane and constitute a mechanism that regulates channel activity.
Biochemical and Biophysical Research Communications 08/2010; 399(3):341-6. · 2.48 Impact Factor
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ABSTRACT: Prokaryotic voltage-gated sodium channels (Na(V)s) are homotetramers and are thought to inactivate through a single mechanism, named C-type inactivation. Here we report the voltage dependence and inactivation rate of the NaChBac channel from Bacillus halodurans, the first identified prokaryotic Na(V), as well as of three new homologues cloned from Bacillus licheniformis (Na(V)BacL), Shewanella putrefaciens (Na(V)SheP), and Roseobacter denitrificans (Na(V)RosD). We found that, although activated by a lower membrane potential, Na(V)BacL inactivates as slowly as NaChBac. Na(V)SheP and Na(V)RosD inactivate faster than NaChBac. Mutational analysis of helix S6 showed that residues corresponding to the "glycine hinge" and "PXP motif" in voltage-gated potassium channels are not obligatory for channel gating in these prokaryotic Na(V)s, but mutations in the regions changed the inactivation rates. Mutation of the region corresponding to the glycine hinge in Na(V)BacL (A214G), Na(V)SheP (A216G), and NaChBac (G219A) accelerated inactivation in these channels, whereas mutation of glycine to alanine in the lower part of helix S6 in NaChBac (G229A), Na(V)BacL (G224A), and Na(V)RosD (G217A) reduced the inactivation rate. These results imply that activation gating in prokaryotic Na(V)s does not require gating motifs and that the residues of helix S6 affect C-type inactivation rates in these channels.
Journal of Biological Chemistry 12/2009; 285(6):3685-94. · 4.77 Impact Factor
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ABSTRACT: The recently developed technique of reductive amination, followed by gold labeling, was applied to visualize the reducing ends of cellulose microcrystals from cellulose I, cellulose II, and cellulose III(I). In these crystals, which were also characterized by electron diffraction, the labeling proved that the chains were organized in a parallel fashion in cellulose I from ramie and Valonia and also in cellulose III(I) from Valonia. In microcrystals of cellulose II from mercerized ramie, the labeling method showed that the chains were packed into an antiparallel mode. These results are discussed in terms of the fine structure of cellulose I where neighboring microfibrils of opposite polarity are visualized. The mercerization process whereby cellulose I is converted into cellulose II is therefore best described in terms of an intermingling of the cellulose chains from neighboring microfibrils of opposite polarity. As opposed to the case of mercerization the conversion of cellulose I into cellulose III(I) does not require the participation of neighboring microfibrils since the crystalline domains are converted individually.
Biomacromolecules 02/2006; 7(1):274-80. · 5.48 Impact Factor
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ABSTRACT: The molecular directionality of chitin biosynthesis was investigated by transmission electron microscopy (TEM) using electron crystallography methods applied to reducing-end-labelled beta-chitin microcrystals from vestimentiferan Lamellibrachia satsuma tubes and nascent beta-chitin microfibrils from the diatom Thalassiosira weissflogii. The data allowed confirmation that the microfibrils were extruded with their reducing end away from the biosynthetic loci, an orientation consistent only with elongation through polymerization at the non-reducing end of the growing chains. Such a chain-extension mechanism, which has also been demonstrated for cellulose and hyaluronan, appears to be general for glycosyltransferases that belong to the GT2 (glycosyl transferase 2) family. The data also allowed confirmation that in beta-chitin the chains are crystallized in a 'parallel-up' mode, in contrast with hypotheses proposed in previous reports.
Biochemical Journal 10/2003; 374(Pt 3):755-60. · 4.90 Impact Factor
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ABSTRACT: A novel procedure for labelling the molecular ends of beta-chitin crystals has been established. By introducing a hydrazide derivative of biotin at the reducing end of a chitin chain, followed by a specific interaction between biotin and streptavidin coupled with a colloidal gold particle, the chain directionality of beta-chitin microcrystals could be directly visualized by transmission electron microscopy. This method allowed to certify the parallelism of the chitin chains in the beta-chitin microcrystals, and also to label the reducing tips of beta-chitin microcrystals degraded by Bacillus circulans chitinase A1. With these substrates, the labelling occurred only at their tapered tip, which indicates that the digestion of these crystals proceeded from their reducing end. The generalization of this new labelling method to other polysaccharide crystals is discussed.
FEBS Letters 02/2002; 510(3):201-5. · 3.54 Impact Factor
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ABSTRACT: The in-depth structural analysis of spherulites of artificial cellulose, formed by the enzymatic polymerization of cellobiosyl fluoride, is reported. Both positive- and negative-type spherulites were observed by polarization optical microscopy. Scanning electron microscopy revealed that the negative spherulites have a three-dimensional round shape, where platelike single crystals of the artificial cellulose originate radially from the center. Transmission electron microscopy confirmed that the cellulose chains orient vertically in the platelike crystals, explaining the observation of negative-type spherulites by polarization optical microscopy. Cellulose spherulites can be formed easily via the enzymatic polymerization process, and the occurrence of both positive- and negative-type spherulites is proposed, due to two independent polymerization mechanisms.
Biomacromolecules 03/2000; 1(2). · 5.48 Impact Factor
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ABSTRACT: Ultrastructural localization of cellulose Iα and Iβ allomorphs in one microfibril from algal sources was investigated using electron microdiffraction. Both cellulose Iα and Iβ were characterized as one-chain triclinic and two-chain monoclinic unit cells, respectively, in agreement with previous studies. These two structures coexisted in each microfibril, alternating either longitudinally or laterally. The transition zone between the two phases was found to be the interface between adjacent H-bonded molecular sheets (i.e., 0.39-nm lattice planes).
08/1998;
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ABSTRACT: Crystalline features of cellulose microfibrils in the cell walls of Glaucocystis (Glaucophyta) were studied by combined spectroscopy and diffraction techniques, and the results were compared with those of Oocystis (Chlorophyta). Although these algae are grouped into two different classes, by the composition of their chloroplasts for instance, their cell walls are quite similar in size and morphology. The most striking features of their cellulose crystallites are that they have the highest cellulose Iα contents reported to date. In particular, the Iα fraction of cellulose from Glaucocystis was found to be as high as 90% from 13C NMR analysis. The mode of preferential orientation of cellulose crystallites in their cell walls is also interesting; equatorial 0.53-nm lattice planes were oriented parallel to the cell surface in the case of Glaucocystis, while the 0.62-nm planes were parallel to the Oocystis cell surface. Such a structural variation provides another link to the evolution of cellulose structure, biosynthesis, and its biocrystallization mechanism.
Journal of Structural Biology.
