白喉毒素-血管内皮生长因子突变体融合毒素的研究

本研究旨在利用突变后的血管内皮生长因子（VEGF）与白喉毒素（DT）构建靶向毒素，使其特异性地杀伤肿瘤血管，达到阻断血管增生，切断肿瘤的血液供应的目的。首先通过基因工程技术从白喉杆菌中提取基因组DNA，扩增出其中的DTC区、T区基因。并运用不同的突变方法，构建了四个VEGF的突变体。即VEGF R82A，K84A，H86A突变，VEGF D63A，E64A，E67A突变，和截去了肝素结合区和NP-l（neuropilin-l）受体结合区的VEGF R82A，K84A，H86A突变和VEGF D63A，E64A，E67A突变。以这些突变体的编码基因与DT的T区、C区基因制成四个融合基因，并在大肠杆菌中表达、纯化、复性。以相似条件下制备的不加VEGF突变体的DTT区、C区蛋白为阴性对照，以VEGFR-l＋和VEGFR-2＋的Lovo细胞和VEGFR-l＋VEGFR-2-的SMMC-7721细胞做细胞实验。制成的四个融合毒素中有三个通过细胞学检验，具有靶向杀伤作用。特异性结合VEGFR-1受体的蛋白PmV8D、PsmVSD既可以抑制VEGFR-1+、VEGFR-2-的肝癌细胞SMMC-7721又可以抑制VEGFR-1+、VEGFR-2＋的结肠癌细胞Lovo。而不带VEGF突变体的DT391无抑制作用。特异性结合VEGFRZ受体的蛋白PmV6D可以抑制VEGFR-1+、VEGFR-2＋结肠癌细胞Lovo的生长，但不可以抑制VEGFR-1＋，VEGFR-2-的肝癌细胞SMMC-7721。受体结合力过弱的PsmV6D不对细胞的生长产生任何影响。

Genomic cloning, expression and recombinant protein purification of alpha and beta subunits of the allophycocyanin gene (apc) from the cyanobacterium Anacystis nidulans UTEX 625

A genomic fragment encoding alpha(APC) and beta(APC) (i.e., alpha and beta units of the allophycocyanin, APC) from Anacystis nidulans UTEX 625 was cloned and sequenced. This fragment, containing a non-coding sequence of 56 nucleotides in between, was then subcloned into the expression vector pMal-c2 downstream from and in frame with the malE gene of E. coli encoding MBP ( maltose binding protein). The fusion protein was purified by amylose affinity chromatography and cleaved by coagulation factor Xa. alpha(APC) and beta(APC) were then separated from MBP and MBP fusion proteins, respectively, and concentrated by membrane centrifugation. The study provides a method to produce recombinant allophycocyanin subunits for biomedical and biotechnological applications.

Cloning and recombinant expression of a crustin-like gene from Chinese shrimp, Fenneropenaeus chinensis

Antimicrobial peptides or proteins (AMPs) are proved to be one of the most important humoral factors to resist pathogen infection. As an antimicrobial protein, crustin had been described in invertebrates as a component of the innate immune system. A crustin-like gene (CruFc) was cloned from haemocytes of Chinese shrimp Fenneropenaeus chinensis by 3' and 5'-RACE PCR. The full-length cDNA consists of 523 with 405 bp open reading frame encoding 134 amino acids and the deduced peptide contains a putative signal peptide of 17 amino acids. The sequence also contains a whey-acidic protein (WAP) domain at the C-terminal. Transcripts of CruFc were mainly detected in haemocytes and gill by RT-PCR analysis. In addition, another full-length cDNA named CshFc was also cloned from haemocytes of Chinese shrimp and its inferred amino acid sequence lacks the WAP-type 'four-disulfide core' domain. The fusion proteins containing CruFc and CshFc were, respectively, produced and the antimicrobial assays revealed that the recombinant CruFc could inhibit the growth of grain-positive bacteria in vitro but the recombinant CshFc could not inhibit at the same conditions. The difference of antimicrobial activity between recombinant CruFc and CshFc provides the evidence that the four-disulfide core domain of crustin may play an important role in its biological function. (c) 2006 Elsevier B.V. All rights reserved.

Proteomic Analysis of Proteins Involved in Spermiogenesis in Mouse

Spermiogenesis is a unique process in mammals during which haploid round spermatids mature into spermatozoa in the testis. Its successful completion is necessary for fertilization and its malfunction is an important cause of male infertility. Here, we report the high-confidence identification of 2116 proteins in mouse haploid germ cells undergoing spermiogenesis: 299 of these were testis-specific and 155 were novel. Analysis of these proteins showed many proteins possibly functioning in unique processes of spermiogenesis. Of the 84 proteins annotated to be involved in vesicle-related events, VAMP4 was shown to be important for acrosome biogenesis by in vivo knockdown experiments. Knockdown of VAMP4 caused defects of acrosomal vesicle fusion and significantly increased head abnormalities in spermatids from testis and sperm from the cauda epididymis. Analysis of chromosomal distribution of the haploid genes showed underrepresentation on the X chromosome and overrepresentation on chromosome 11, which were due to meiotic sex chromosome inactivation and expansion of testis-expressed gene families, respectively. Comparison with transcriptional data showed translational regulation during spermiogenesis. This characterization of proteins involved in spermiogenesis provides an inventory of proteins useful for understanding the mechanisms of male infertility and may provide candidates for drug targets for male contraception and male infertility.

Silicon Surface Modification with a Mixed Silanes Layer to Immobilize Proteins for Biosensor with Imaging Ellipsometry

One kind of surface modification method on silicon wafer was presented in this paper. A mixed silanes layer was used to modify silicon surface and rendered the surface medium hydrophobic. The mixed silanes layer contained two kinds of compounds, aminopropyltriethoxysilane (APTES) and methyltriethoxysilane (NITES). A few of APTES molecules in the layer was used to immobilize covalently human immunoglobulin G (IgG) on the silicon surface. The human IgG molecules immobilized covalently on the modified surface could retain their structures well and bind more antibody molecules than that on silicon surface modified with only APTES. This kind of surface modification method effectively improved the sensitivity of the biosensor with imaging ellipsometry.

Turbulent Modeling of ABL over Wavy Water Surface

A new model accounting for both turbulence and sea state effects has been proposed in this paper to describe momentum, heat and moisture exchanges through air-sea interface. While long wave components mainly change air flow profile, short wave components

Optimization of 2-stage gas gun for fusion refueling pellet injection with consideration of real effects

Analytic expression of pellet acceleration by constant base pressure with consideration of gas-wall friction, heat transfer and viscous dissipation that important for high speed injection is obtained. The process of compression stage is formulated by a set of governing equations and can be numerically integrated. Excellent confirmation with experiments is obtained and the ways to optimum match the compression stage with the launch stage are suggested.

Fusion energy-production from a deuterium-tritium plasma in the jet tokamak

Describes a series of experiments in the Joint European Torus (JET), culminating in the first tokamak discharges in deuterium-tritium fuelled mixture. The experiments were undertaken within limits imposed by restrictions on vessel activation and tritium usage. The objectives were: (i) to produce more than one megawatt of fusion power in a controlled way; (ii) to validate transport codes and provide a basis for accurately predicting the performance of deuterium-tritium plasmas from measurements made in deuterium plasmas; (iii) to determine tritium retention in the torus systems and to establish the effectiveness of discharge cleaning techniques for tritium removal; (iv) to demonstrate the technology related to tritium usage; and (v) to establish safe procedures for handling tritium in compliance with the regulatory requirements. A single-null X-point magnetic configuration, diverted onto the upper carbon target, with reversed toroidal magnetic field was chosen. Deuterium plasmas were heated by high power, long duration deuterium neutral beams from fourteen sources and fuelled also by up to two neutral beam sources injecting tritium. The results from three of these high performance hot ion H-mode discharges are described: a high performance pure deuterium discharge; a deuterium-tritium discharge with a 1% mixture of tritium fed to one neutral beam source; and a deuterium-tritium discharge with 100% tritium fed to two neutral beam sources. The TRANSP code was used to check the internal consistency of the measured data and to determine the origin of the measured neutron fluxes. In the best deuterium-tritium discharge, the tritium concentration was about 11% at the time of peak performance, when the total neutron emission rate was 6.0 × 10¹⁷ neutrons/s. The integrated total neutron yield over the high power phase, which lasted about 2 s, was 7.2 × 10¹⁷ neutrons, with an accuracy of ±7%. The actual fusion amplification factor, Q_DT was about 0.15

Generating artificial homologous proteins according to the representative family character in molecular mechanics properties - an attempt in validating an underlying rule of protein evolution

The molecular mechanics property is the foundation of many characters of proteins. Based on intramolecular hydrophobic force network, the representative family character underlying a protein’s mechanics property is described by a simple two-letter scheme. The tendency of a sequence to become a member of a protein family is scored according to this mathematical representation. Remote homologs of the WW-domain family could be easily designed using such a mechanistic signature of protein homology. Experimental validation showed that nearly all artificial homologs have the representative folding and bioactivity of their assigned family. Since the molecular mechanics property is the only consideration in this study, the results indicate its possible role in the generation of new members of a protein family during evolution.

Laser-confined fusion

An approach for producing a large quantity of neutrons is proposed. It involves compression of a fuel foil and confinement of the resulting plasma between two intense laser pulses. It is shown that two circularly polarized laser pulses of amplitude a=7 illuminating a deuterium-tritium foil of areal density 3.3 X 10(18) cm(-2) can produce about 4.2 X 10(6) neutrons per joule of the input laser energy.