Analysis of alcohol oxidase isozymes in gene-disrupted strains of methylotrophic yeast Pichia methanolica

A cell-free extract of methanol-grown Pichia methanolica cells was found to contain nine alcohol oxidase (AOD) isozymes by active staining of a native polyacrylamide electrophoresis gel. Our previous study revealed that AOD in P. methanolica was encoded by two genes, MOD1 and MOD2, and the results of an experiment involving Candida boidinii as an expression host suggested that the AOD isozymes observed in P. methanolica were due to random association of Mod1p and Mod2p into an active octamer [Nakagawa et al., Yeast, 15, 1223-1230 (1999)]. This study was conducted using P. methanolica MOD1- and/or MOD2-gene disrupted strains to confirm a previous hypothesis. While the cell-free extract of the wild-type strain gave nine ladder bands, the mod1delta and mod2delta strains gave a single active AOD band corresponding to the mobilities of Mod2p and Mod1p on a native electrophoresis gel, respectively. The cell-free extract of glyceorl-grown wild-type cells gave a single band corresponding to Mod1p, showing that only MOD1 is expressed in glycerol-grown cells. While the expression of both MOD1 and MOD2 was induced by methanol, this finding and our previous observations indicated that the expression of MOD1 and MOD2 was controlled by a distinct regulatory mechanism in P. methanolica.     

On one- and two-parameter analyses of short fatigue crack growth

Recent data on short fatigue crack growth in two cast and hot isostatically pressed (hipped) aluminum alloys obtained by Shyam, Allison and Jones have been analyzed in terms of a previously proposed one-parameter short crack model which includes consideration of elastic–plastic effects, the Kitagawa effect and the development of crack closure in the wake of a newly formed crack. The material constants obtained in a prior investigation of short crack growth behavior in a cast aluminum alloy tested under fully reversed loading were used as a basis for the present analysis. The predicted rates of fatigue crack propagation are in accord with the experimental results. In the discussion, aspects of the two-parameter approach presented by Shyam et al. are compared with those of the one-parameter method of analysis used herein.     

Cover Picture: Assembly and Photoinduced Organization of Mono‐ and Oligopeptide Molecules Containing an Azobenzene Moiety (Adv. Funct. Mater. 9/2007)

An illustration of the phototriggered organization and dispersion of tri‐peptide‐substituted azobenzene derivatives, as reported on p. 1507 by Yoko Matsuzawa and co‐workers. The tri‐peptide units form a beta‐sheet structure through hydrogen bonding to assemble a fibrous network system. Cis–trans photoisomerization of the azobenzene moiety leads to a large polarity change of the component molecule to break and re‐form hydrogen bonds between the tri‐peptide units. Assembled systems consisting of an azobenzene moiety as the photofunctional component and valyl units as the network backbone are investigated. The molecular ordering of these assemblies is examined by spectroscopy and theoretical calculations. The number of valyl units greatly influences the molecular order in the organized systems. Only N‐(L ‐valyl‐L ‐valyl‐L ‐valyl)azobenzene‐4‐carboxamide ( 3 ) forms a complete β‐sheet structure in this artificial assembly. Upon photoirradiation, the azobenzene moieties isomerize completely, revealing the reversibility in the structural organization through the flexibility of the β‐sheet network in this system.     

Study of local intracellular signals regulating axonal morphogenesis using a microfluidic device

The establishment and maintenance of axonal patterning is crucial for neuronal function. To identify the molecular systems that operate locally to control axonal structure, it is important to manipulate molecular functions in restricted subcellular areas for a long period of time. Microfluidic devices can be powerful tools for such purposes. In this study, we demonstrate the application of a microfluidic device to clarify the function of local Ca²⁺ signals in axons. Membrane depolarization significantly induced axonal branch-extension in cultured cerebellar granule neurons (CGNs). Local application of nifedipine using a polydimethylsiloxane (PDMS)-based microfluidic device demonstrated that Ca²⁺ entry from the axonal region via L-type voltage-dependent calcium channels (L-VDCC) is required for branch extension. Furthermore, we developed a method for locally controlling protein levels by combining genetic techniques and use of a microfluidic culture system. A vector for enhanced green fluorescent protein (EGFP) fused to a destabilizing domain derived from E. coli dihydrofolate reductase (ecDHFR) is introduced in neurons by electroporation. By local application of the DHFR ligand, trimethoprim (TMP) using a microfluidic device, we were able to manipulate differentially the level of fusion protein between axons and somatodendrites. The present study revealed the effectiveness of microfluidic devices to address fundamental biological issues at subcellular levels, and the possibility of their development in combination with molecular techniques.     

Photorefractive device using self-assembled monolayer coated indium-tin-oxide electrodes

Photorefractive (PR) performances of methyl-substituted poly(triarylamine) (PTAA)-based PR device were demonstrated using chemically modified electrodes (CMEs) of self-assembled monolayer (SAM) coated indium-tin-oxide (ITO) electrodes. The SAM-ITO electrodes successfully suppressed dark current which blocks the formation of space-charge field and also causes the dielectric breakdown. The PR device consisted of composite of PTAA, 4-azacycloheptylbenzylidenemalononitrile (7-DCST), N-ethylcarbazole (ECz), and [6,6]-phenyl-C61-butyric acid methyl ester (PCBM) sandwiched between the SAM-ITO electrodes. The PR devices showed the PR performances: optical gain Γ, refractive index Δn, diffraction efficiency η, response time τ, sensitivity S, phase shift Φ, trap-limited field E_q, number density of traps N_T, and space-charge field E_SC. The remarkable response time of 11.3 ms was achieved at the low electric field of 20 V μm⁻¹, which was comparable to the response time of high-definition television (HDTV) quality of 16 ms. Our approach will widen the usage of higher mobility materials to photorefractive field and give us more favorable materials to achieve the best performance of photorefractivity in the future.     

Numerical Evaluation of Toughening by Crack-Face Grain Interlocking in Self-Reinforced Ceramics

Crack bridging associated with the pull-out process of interlocking grains in self-reinforced ceramic materials is studied through a micromechanical simulation. The pullout of a single inclined grain is modeled via the numerical solution of a general contact problem. The bridging-force versus crack-opening-distance curve indicates a nonlinear, springlike response for the pullout of interlocking grains. The sliding friction along the debonded interface, induced by highly localized contact stresses, dominates the total bridging force. The bridging force increases with grain inclination until eventual bridge failure. The pullout of misaligned grains mainly affects short-crack toughening, with a rising R -curve, whereas aligned grains contribute to long-crack toughening. The residual stresses of the thermal expansion anisotropy play a minor role in the pull-out process of grain interlocking and the resultant toughening. The proposed mechanism is operative in both single-phase and composite ceramics in which pullout of elongated grains/reinforcements occurs.     

Synthesis and characterization of nanocrystalline Mg2CoH5 obtained by mechanical alloying

The stoichiometric mixture of 2MgH₂ + Co was ball milled under a hydrogen atmosphere to synthesize nanocrystalline metal hydride Mg₂CoH₅. Upon milling, the mixture was analyzed by X-ray powder diffraction (XRD) and thermal methods employing the techniques of differential scanning calorimetry (DSC), thermogravimetry (TG) and differential thermal analysis (DTA). Hydrogen absorption and desorption measured by pressure-composition-temperature (P-C-T) curves indicated that the capacity loss was small after 20 consecutive cycling tests. The enthalpies associated with hydride formation and decomposition were measured to be –69.5 and –83.2 kJ mol^–1 H₂, respectively. At the temperatures of this study (553 to 653 K), hysteresis decreases with increasing temperature.     

TEM,XPS and SIMS Analyzes on Grain Boundary of Lanthanum Chromites

The morphological characteristics, chromium valence state, and cation transport in the vicinity of grain boundary in La_1-xCa_xCrO₃ were investigated by using TEM/EDS, XPS and SIMS techniques. The width of grain boundary was around 1 nm where anomalous enrichment of calcium was observed. Higher valence state of chromium such as Cr⁶⁺(d⁰) was detected in the grain boundaries whereas Cr³⁺(d³) and Cr⁴⁺(d²) were dominant in the bulk. Very fast interdiffusion of alkaline earths was observed in the Sr²⁺-La_0.75Ca_0.25CrO₃ system. All observed phenomena were correlated by assuming the A-site vacancy which may be induced by the formation of Cr⁶⁺ at grain boundaries.     

Debye-Waller parameters of diatomic crystals

The Debye-Waller parameters, the effective valuesB _eff, the dynamic component,B _d, and the static component,B _s, of each constituent atom and also of the bulk were measured for some diatomic crystals with NaCl- and zinc blende-type structures. The dominant effect of heavy constituent atoms on the dynamic component of bulk crystals was found, and the mean square displacement in bulk crystals was approximated well by the atomic mass averaged mean square displacements of the constituent atoms. A remarkable effect of structural defects onB _eff, particularly onB _s, was observed in NiO samples with different degrees of non-stoichiometry. The different extents of the SiC-forming reaction between solid carbon and fused silicon resulted in quite differentB _eff values; the larger theB _eff value the smaller the extent of the reaction.