Trypsiligase-Catalyzed Labeling of Proteins on Living Cells

Fluorescent fusion proteins are powerful tools for studying biological processes in living cells, but universal application is limited due to the voluminous size of those tags, which might have an impact on the folding, localization or even the biological function of the target protein. The designed biocatalyst trypsiligase enables site-directed linkage of small-sized fluorescence dyes on the N terminus of integral target proteins located in the outer membrane of living cells through a stable native peptide bond. The function of the approach was tested by using the examples of covalent derivatization of the transmembrane proteins CD147 as well as the EGF receptor, both presented on human HeLa cells. Specific trypsiligase recognition of the site of linkage was mediated by the dipeptide sequence Arg-His added to the proteins’ native N termini, pointing outside the cell membrane. The labeling procedure takes only about 5 minutes, as demonstrated for couplings of the fluorescence dye tetramethyl rhodamine and the affinity label biotin as well.     

Fabrication and joining mechanism of Nano-Cu/Si3N4 ceramic substrates

A novel method for fabricating a nano-Cu/Si₃N₄ ceramic substrate is proposed. The nano-Cu/Si₃N₄ ceramic substrate is first fabricated using spark plasma sintering (SPS) with the addition of nanoscale multilayer films (Ti/TiN/Ti/TiN/Ti) as transition layers. The microstructures of the nano-Cu metal layer and the interface between Cu and Si₃N₄ are investigated. The results show that a higher SPS temperature increases the grain size of the nano-Cu metal layer and affects the hardness. The microstructure of the transition layer evolves significantly after SPS. Ti in the transition layer can react with Si₃N₄ and with nano-Cu to form interfacial reaction layers of TiN and Ti–Cu, respectively; these ensure stronger bonding between nano-Cu and Si₃N₄. Higher SPS temperatures improve the diffusion ability of Ti and Cu, inducing the formation of Ti₃Cu₃O compounds in the nano-Cu metal layer and Ti₂Cu in the transition layer. This study provides an important strategy for designing and constructing a new type of ceramic substrate.     

Nonlinear response in glass fibre non-crimp fabric reinforced vinylester composites

This paper addresses the nonlinear stress-strain response in glass fibre non-crimp fabric reinforced vinylester composite laminates subjected to in-plane tensile loading. The nonlinearity is shown to be a combination of brittle and plastic failure. It is argued that the shift from plastic to brittle behaviour in the vinylester is caused by the state of stress triaxiality caused by the interaction between fibre and vinylester. A model combining damage and plasticity is calibrated and evaluated using data from extensive experimental testing. The onset of damage is predicted using the Puck failure criterion, and the evolution of damage is calibrated from the observed softening in plies loaded in transverse tension. Shear loading beyond linear elastic response is observed to result in irreversible strains. A yield criterion is implemented for shear deformation. A strain hardening law is fitted to the stress-strain response observed in shear loaded plies. Experimental results from a selection of laminates with different layups are used to verify the numerical models. A complete set of model parameters for predicting elastic behaviour, strength and post failure softening is presented for glass fibre non-crimped fabric reinforced vinylester. The predicted behaviour from using these model parameters are shown to be in good agreement with experimental results.     

Densification mechanism during reactive hot pressing of B4C-ZrO2 mixtures

The densification behaviors of pure B₄C and B₄C-ZrO₂ mixtures were compared during hot pressing. The results showed that in-situ formed ZrB₂ effectively enhanced the densification process of B₄C-ZrO₂ mixtures, more significantly during the intermediate stage. Within the relative density ranging from 0.75 to 0.90, the B₄C-15?wt%ZrO₂ mixture (B15Z) achieved the maximum densification rate as twice much as that of pure B₄C. The stress exponent n>3 indicated plastic deformation was the dominant densification mechanism of B15Z. The viscosities of plastic flow were evaluated using Murray-Rodger-William equation and the viscosity of B15Z was only a quarter of that in pure B₄C. The sintering activation energy was calculated to be 305.9?kJ/mol for pure B₄C and 197?kJ/mol for B15Z, respectively. It was proposed that the lower viscosity of plastic flow and activation energy accelerated the sliding and propagating motions of plastic flow, by which underlain the enhanced densification behaviors of B₄C-ZrO₂ mixtures.     

Design and Characterization of a New pVII Combinatorial Phage Display Peptide Library for Protease Substrate Mining Using Factor VII Activating Protease (FSAP) as Model

We describe a novel, easy and efficient combinatorial phage display peptide substrate-mining method to map the substrate specificity of proteases. The peptide library is displayed on the pVII capsid of the M13 bacteriophage, which renders pIII necessary for infectivity and efficient retrieval, in an unmodified state. As capture module, the 3XFLAG was chosen due to its very high binding efficiency to anti-FLAG mAbs and its independency of any post-translational modification. This library was tested with Factor-VII activating protease (WT-FSAP) and its single-nucleotide polymorphism variant Marburg-I (MI)-FSAP. The WT-FSAP results confirmed the previously reported Arg/Lys centered FSAP cleavage site consensus as dominant, as well as reinforcing MI-FSAP as a loss-of-function mutant. Surprisingly, rare substrate clones devoid of basic amino acids were also identified. Indeed one of these peptides was cleaved as free peptide, thus suggesting a broader range of WT-FSAP substrates than previously anticipated.     

Effect of grinding-induced cyclic heating on the hardened layer generation in the plunge grinding of a cylindrical component

This paper discusses the effects of the grinding-induced cyclic heating on the properties of the hardened layer in a plunge cylindrical grinding process on the high strength steel EN26. It was found that a multi-pass grinding brings about a uniform and continuous hardened layer along the circumference of the cylindrical workpiece. An increase of the number of grinding passes, leads to a thicker layer of hardening, a larger compressive residual stress and a deeper plastic deformation zone. Within the plastic deformation zone, the martensitic grains are refined by the thermo-mechanical loading, giving rise to a hardness of 12.5% higher than that from a conventional martensitic transformation. The coupled effects of heat accumulation and wheel wear in the multi-pass grinding are the main causes for the thickening of the hardened layer. A too small infeed per workpiece revolution would result in insufficient grinding heat, and in turn, bring about an undesirable tempered hardened layer and a reduction of its hardness.     

Stress-strain relations in elastoplastic solids with Dugdale-type cracks

Mechanical behavior of a two-dimensional elastoplastic solid with rectilinear cracks is investigated. Plastic strip model is used to reduce plasticity problem to the equivalent linear elasticity formulation. Two realizations of the mixed mode plastic strip model are considered: in-line plastic strips as proposed by Becker and Gross [Int. J. Fract. 37 (1988) 163], and inclined plastic strips of Panasyuk and Savruk [Appl. Mech. Rev. 47 (1994) 151]. The effective mechanical response predictions are based on the procedure presented in Kachanov et al., [Appl. Mech. Rev. 47 (1994) 151]. Stress-strain relations are obtained for parallel and randomly oriented non-interacting cracks. Results are compared with known elastic solutions.     

The influence of plastic properties on chip formation

A two-dimensional finite-element model of the chip formation process is used to study the influence of the material law determining plastic flow on chip formation of titanium alloys at high cutting speeds. For simplicity, friction and thermo-mechanical properties of the tool are neglected in the simulations. Titanium chips are strongly segmented and therefore especially suitable to study the role of segmentation for the cutting force and other cutting parameters. Of special interest is the influence of the thermal softening parameter in the material law. Increasing thermal softening leads to a drastic decrease in the cutting force and a corresponding increase in chip segmentation. A variation of the hardening exponent is also performed. It is shown that the simulation results can be understood by using adiabatic flow stress curves. The variation of the hardening exponent leads to strongly differing chip shapes, although the cutting force stays nearly constant. This shows that mean cutting forces should not be used as simulation verification parameters.     

二氧化锡溅射工艺研究

SnO2 是最早使用也是目前使用最广泛的一种气敏材料 ,使用该材料设计制作的气敏传感器具有许多优点。在简要介绍溅射镀膜的成膜过程和特点的基础上 ,着重介绍了SnO2 膜的制备流程 ,分析了功率和温度变化对成膜质量的影响     

塑料模现状及发展趋势

我国塑料模的发展,极其迅速,现已具相当规模。无论是设计水平、加工制造技术,还是 CAD 技术都有了明显提高。本文简明扼要地介绍了国内外塑料模的现状、发展趋势、市场预测,以及“九五”国家规划目标与对策。