Expression of human β-subunit nerve growth factor (hNGFB) in yeast Pichia pastoris and E. coli

The gene hNGFB encoding the β subunit of human nerve growth factor (hNGF) was cloned intoP. pastoris secretive expression vector pHIL-S1 andE. coli expression vector pET-15b. The recombinant hNGFB vectors pSNGF and pET15b-NGF were transformed intoP. pastoris host cell GS115 (Mut⁺, His⁻) andE. coli strain BL21 (DE3) respectively. Expression and secretion of hNGFB inP. pastoris was attempted under the direction of the AOX1 promoter and PHO1 signal sequence. The positive colonies growing on medium without histidine were further selected by PCR. The yield of rehNGFB in GS115 was about 14.4% of total cellular secretive protein. The secreted protein was immunological active on Western blotting with rabbit anti-mNGFB antibodies. The fusion protein yield of rehNGFB inE. coli BL21 (DE3) was about 10.3% of total cellular protein after IPTG induction. Western blot detection showed its immunological activity.     

亲和层析的定量分析

亲和层析技术广泛地用于多糖蛋白质，核酸，激素，维生素和细胞等的分离与纯化。近年来，亲和层析技术逐渐被用于定量分析，并且在生物和临床医学的研究的研究及对某些疾病进行检验和诊断中获得了成功。     

横观各向同性矩形板的非经典稳定问题

针对四边简支的横观各向同性矩形板单向受非均匀对称压力作用的稳定问题给出了三维分析方法.首先通过引入位移函数和应力函数,从三维基本方程出发构造了两类相互独立的状态空间方程.其次结合层合近似理论,求解了对应非经典稳定问题的状态空间方程.进而利用子层间的连续条件和矩形板上下表面的自由条件,导出了两类独立的特征方程.     

Formation of a new fluorescence of human neuronal tau protein

A new fluorescence at 333 nm formed when the solution of human neuronal tau protein was incubated at 37°C. The maximum excitation was at 230 and 280 nm respectively with the quantum yield of about 0.02. It took about 30 h for the complete formation of the new fluorescence and the light scattering increased with the formation procedure. It suggests that the fluorescence resulted from the aggregation of tau molecules in solution.     

d-Ribosylated Tau forms globular aggregates with high cytotoxicity

Although the glycation of Tau that is involved in paired helical filament formation in Alzheimer’s disease has been widely studied, little attention has been paid to the role of d-ribose in the glycation of Tau. Here, we show that Tau is rapidly glycated in the presence of d-ribose, resulting in oligomerization and polymerization. Glycated derivatives appeared after 24 h incubation. Western blotting indicated the formation of advanced glycation end-products (AGEs) during initial stages of glycation. Thioflavin T-positive (ThT-positive) aggregations that appeared from day 4 indicated the globular-like features. Atomic force microscopy revealed that the surface morphology of ribosylated Tau40 was globular-like. Kinetic studies suggested that d-ribosylated Tau is slowly oligomerized and rapidly polymerized with ThT-positive features. Moreover, d-ribosylated Tau aggregates were highly toxic to SHSY5Y cells and resulted in both apoptosis and necrosis. This work has demonstrated that d-ribose reacted with Tau protein rapidly, producing ThT-positive aggregations which had high cytotoxicity. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

编制县级土地利用总体规划图若干问题探讨

县土地利用总体规划图是土地利用规划的主要研究成果之一.该文在实践的基础上就编制规划图中的若干问题进行了探讨.