藏猪肠道不同部位CAT1、EAAC1和PepT1 mRNA的特异性分布

选取体重接近且健康的28日龄哺乳仔藏猪6头,屠宰后分离肠道(十二指肠、空肠前段、空肠后段和回肠),运用RFQ-PCR技术对藏猪不同肠段小肽转运载体PepT1、酸性氨基酸转运载体EAAC1和碱性氨基酸转运载体CAT1 mRNA的组织分布进行研究.结果表明:空肠后段PepT1 mRNA表达丰度最高,而回肠的表达量最低;从小肠近端到远端EAAC1 mRNA的表达丰度在小肠的表达呈升高趋势,即回肠表达最高,空肠后段次之,但与十二指肠、空肠前段差异不显著(P>0.05);CAT1 mRNA不同肠段表达格局与EAAC1相似,回肠中CAT1 mRNA表达丰度最高,显著高于十二指肠(P<0.05),但空肠前、后段的差异不显著(P>0.05).     

酵母Mn-transporter基因研究进展

在酵母中有两个锰离子的转运系统 :即高亲和力系统和低亲和力系统 .本文简要介绍了酵母中与锰离子转运有关的基因 :SMF1、BSD2、ATX2、CCC1、PMR1和MNR1及它们所编码蛋白的大小、细胞定位、跨膜域及他们在锰转运中的功能 .文中还比较了几种蛋白之间的关系 .     

饥饿再投喂对鳜肌FSRP-1、FSRP-3和肠道PepT1基因表达的影响

为探讨饥饿再投喂对鳜肌FSRP-1、FSRP-3和肠道中PepT1基因表达的影响,本研究采用qRT-PCR技术,测定饥饿7d再饱食一餐后0、1、3、6、12、24、48、96h条件下鳜肌中FSRP-1、FSRP-3和肠道中PepT1基因表达水平的变化,研究结果表明:FSRP-1基因表达在投喂后12h内显著升高(P0.05),12-48h维持在较高表达水平,48h后显著降低(P0.05);FSRP-3基因表达在投喂后1h内显著降低(P0.05),12h达到峰值且表达量是1h的5倍,48h后显著降低(P0.05);PepT1基因表达在投喂后6h内显著降低(P0.05),48h时表达量最高。FSRP-1、FSRP-3和PepT1是与鱼类生长相关的重要基因,受饥饿再投喂的影响较大。     

小拟南芥转运蛋白基因及NHX2基因的克隆与表达

转运蛋白(transport protein)是膜蛋白的一大类,介导生物膜内外的化学物质及信号的交换。植物体内存在多个与Na~+转运相关的蛋白,其中液泡膜Na~+/H~+逆向转运蛋白(Vacuolar Na~+/H~+antiporter,NHX)在离子稳态和提高植物耐盐性方面发挥着重要作用。为了深入了解转运蛋白基因在短命植物小拟南芥(Arabidopsis pumila)耐盐方面的作用,本研究首先基于小拟南芥响应高盐胁迫叶片转录组数据筛选出1157个转运蛋白基因,按功能分为Na~+转运蛋白,K~+转运蛋白,Ca~(2+)转运蛋白,ABC转运蛋白以及糖转运蛋白等,其中功能注释为Na~+转运蛋白的基因有24个。K均值(K-means)聚类分析结果显示,1157个转运蛋白基因分布于20个K subcluster,其中在K6、K9、K15子聚类中的基因数量分布较多,分别为172、193、190个。在分布于K6子聚类的Na~+转运蛋白基因中,有一个编码NHX2蛋白的基因经盐胁迫处理后明显上调表达。采用RT-PCR克隆了Ap NHX2基因,Ap NHX2开放阅读框1626 bp,编码541个氨基酸。Ap NHX2蛋白是一个典型的跨膜转运蛋白,具有12个跨膜结构区。系统进化分析表明Ap NHX2与拟南芥At NHX2亲缘关系最近。实时荧光定量PCR分析显示,Ap NHX2基因在小拟南芥各组织中均有表达,但在花中表达量最高。为进一步研究该基因的功能,构建了过量表达载体35S∶Ap NHX2并转化农杆菌GV3101。本研究为进一步阐述转运蛋白基因在小拟南芥响应盐胁迫中的功能机制奠定了基础。     

凋亡素融合蛋白的基因克隆、表达与体外活性分析

来源于HIV-1病毒(47~57位氨基酸)的TAT小肽具有跨膜功能,可以将外源蛋白进行跨膜转运。通过聚合酶链反应(PCR)的方法扩增了TAT-凋亡蛋白序列,与载体pET-28b连接后在大肠杆菌BL21(DE3)中获得了高表达,以包涵体形式表达的TAT-凋亡蛋白在变性条件下进行了Ni-NTA纯化,纯化的蛋白经MTT法证明具有诱导HeLa细胞凋亡的能力。     

PEP-3与HBc融合原核表达载体的构建与表达

以可诱导较强免疫反应及呈现高拷贝抗原小肽的乙肝病毒核心蛋白(Hepatitis B Virus Core Protein,HBc)类病毒颗粒(Virus-like Particles,VLPs)作为免疫载体蛋白,通过分子生物学手段,将蓝白班筛选元件Lac Zα插入HBc的主要免疫显性区域,构建获得外源小肽与HBc融合的通用原核表达载体。在此通用载体的基础上,通过PEP-3小肽替换Lac Zα片段,构建了PEP-3与HBc融合原核表达载体,并成功进行了HBc/PEP-3融合蛋白的表达及纯化。该新型通用载体的成功构建,将为小肽疫苗的研究及应用提供有效工具。     

融合聚精氨酸九肽的小热休克蛋白 原核表达及蛋白纯化

目的构建小热休克蛋白-聚精氨酸九肽融合蛋白的原核表达载体,表达纯化融合蛋白.方法在小热休克蛋白表达载体的基础上利用点突变方法引入聚精氨酸九肽对应序列,转入BL21大肠杆菌感受态细胞进行原核表达,利用亲和层析方法对表达蛋白进行纯化.结果成功构建了融合聚精氨酸九肽的HSP16. 5融合蛋白表达载体,并对其在大肠杆菌细胞中的原核表达进行条件优化,研究显示:在诱导剂IPTG浓度为0. 5 mmol、37℃条件下诱导4 h目的蛋白产量较高.结论此实验成功构建了小热休克蛋白-聚精氨酸九肽融合蛋白的原核表达载体,得到了高纯度的小热休克蛋白-聚精氨酸九肽融合蛋白,为进一步研究其功能奠定基础.     

RT-PCR法检测大鼠组织中GLUT8基因表达特异性

采用半定量RT_PCR技术检测青春前期和成年大鼠的主要组织中GLUT8基因mRNA表达水平.实验结果显示,葡萄糖转运蛋白GLUT8基因在成年和青春前期大鼠的睾丸组织中都有高度表达,在心脏和肾脏组织中有少量表达,在肝、脾等组织中的表达是微量的;在成年大鼠睾丸组织中,Leydig细胞、睾丸生精细胞以及附睾的精子细胞中GLUT8基因都有大量的表达,其中以在Leydig细胞中的表达最丰富,而且GLUT8基因在睾丸生精细胞中的表达水平高于附睾的精子细胞.结果说明GLUT8蛋白主要分布在大鼠睾丸组织中,并且主要在Leydig细胞和精子细胞中参与葡萄糖转运和能量代谢与供应.     

酵母Mn—transporter基因研究进展

要酵母中有两个锰离子的转运系统：即高亲和力系统和低亲和力系统。本文简要介绍了酵母中与锰离子转运有关的基因：ＳＭＦ１、ＢＳＤ２、ＡＴＸ２、ＣＣＣ１、ＰＭＲ１和ＭＮＲ１及它们所编码蛋白的大小、细胞定位、跨膜域及他们在锰转运中的功能。文中不比较几中蛋白之间的关系。     

酵母高亲和力铁转运系统主要基因及环境因子对该系统的影响

Fet3基因编码的一种多铜氧化酶和Ftr1基因编码的一种通透酶介导酵母细胞质膜高亲和力的铁吸收,FET3的Fe(Ⅱ）氧化酶活性是酵母高亲和力铁吸收系统必需的,FTR1在铁的跨膜转运中起着直接的作用,其转运底物是Fe(Ⅲ）,O2通过转录因子AFT1的作用和作为FET3的专性底物而对酵母高亲和力的铁吸收系统起着重要的调节作用。     

植物钾离子转运相关蛋白及基因研究进展

本文概述了植物体内与K^ 转运相关的蛋白及其基因，包括通道蛋白(channel protein)和转运体(transporter)及其基因,前者可分为：(1)内向整流K^ 通道(inward-rectifying K^ channel:Kin^ )，(2)外向整流K^ 通道(outward-rectifyjng K^ channel ：Kout^ )；相关基因有AKTI，ANTI，SORK，GORK等．后者分为低亲和K^ 吸收转运体及高亲和K^ 吸收转运体；相关基因有HAK，KUP等．     

Modulation of the neuronal glutamate transporter EAAC1 by the interacting protein GTRAP3-18

Excitatory amino-acid carrier 1 (EAAC1) is a high-affinity Na+-dependent L-glutamate/D,L-aspartate cell-membrane transport protein. It is expressed in brain as well as several non-nervous tissues. In brain, EAAC1 is the primary neuronal glutamate transporter. It has a polarized distribution in cells and mainly functions perisynaptically to transport glutamate from the extracellular environment. In the kidney it is involved in renal acidic amino-acid re-absorption and amino-acid metabolism. Here we describe the identification and characterization of an EAAC1-associated protein, GTRAP3-18. Like EAAC1, GTRAP3-18 is expressed in numerous tissues. It localizes to the cell membrane and cytoplasm, and specifically interacts with carboxy-terminal intracellular domain of EAAC1. Increasing the expression of GTRAP3-18 in cells reduces EAAC1-mediated glutamate transport by lowering substrate affinity. The expression of GTRAP3-18 can be upregulated by retinoic acid, which results in a specific reduction of EAAC1-mediated glutamate transport. These studies show that glutamate transport proteins can be regulated potently and that GTRAP can modulate the transport functions ascribed to EAAC1. GTRAP3-18 may be important in regulating the metabolic function of EAAC1.     

Insulin-regulatable tissues express a unique insulin-sensitive glucose transport protein

At least three different glucose transport systems exist in mammalian cells. These are: (1) the constitutively active, facilitative carrier characteristic of human erythrocytes, Hep G2 (ref. 2) cells and rat brain; (2) the Na-dependent active transporter of kidney and small intestine; and (3) the facilitative carrier of rat liver (B. Thorens and H. F. Lodish, personal communication). A fourth possible glucose transport system is the insulin-dependent carrier that may be specific to muscle and adipose tissue. This transporter resides primarily in an intracellular compartment in resting cells from where it translocates to the cell surface upon cellular insulin exposure. This raises the question of whether hormonal regulation of glucose transport is conferred by virtue of a tissue-specific signalling mechanism or a tissue-specific glucose transporter. Here we present data supporting the latter concept based upon a monoclonal antibody against the fat cell glucose transporter that identifies a unique, insulin-regulatable glucose transport protein in muscle and adipose tissue.     

An efflux transporter of silicon in rice

Silicon is an important nutrient for the optimal growth and sustainable production of rice. Rice accumulates up to 10% silicon in the shoot, and this high accumulation is required to protect the plant from multiple abiotic and biotic stresses. A gene, Lsi1, that encodes a silicon influx transporter has been identified in rice. Here we describe a previously uncharacterized gene, low silicon rice 2 (Lsi2), which has no similarity to Lsi1. This gene is constitutively expressed in the roots. The protein encoded by this gene is localized, like Lsi1, on the plasma membrane of cells in both the exodermis and the endodermis, but in contrast to Lsi1, which is localized on the distal side, Lsi2 is localized on the proximal side of the same cells. Expression of Lsi2 in Xenopus oocytes did not result in influx transport activity for silicon, but preloading of the oocytes with silicon resulted in a release of silicon, indicating that Lsi2 is a silicon efflux transporter. The identification of this silicon transporter revealed a unique mechanism of nutrient transport in plants: having an influx transporter on one side and an efflux transporter on the other side of the cell to permit the effective transcellular transport of the nutrients.     

从七肽噬菌体展示库中筛选蛋白质配基

为了研究噬菌体展示技术的应用，以溶菌酶为靶分子从七肽噬菌体展示库中筛选蛋白质的高亲和力噬菌体配体，所筛选的亲和力最高的噬菌体的ELISA检测值A405nm可达0．634．通过比较亲和性噬菌体外源插入肽的DNA序列，认为基元HWWW是肽段与酶分子发生亲和的必需序列．此外，由于靶分子和高亲和性展示肽的等电点分别为11．2和6．74，因此在亲和环境中携带异种电荷，利于亲和吸附的发生，而此时低亲和性展示肽与靶分子携带同种电荷，阻碍了亲和吸附．同时，高亲和性肽段HWWPAS和与其有较高同源性的肽段HWTWWNL都有适中的疏水性，这有利于肽与靶分子表面的疏水位点相互作用从而产生亲和吸附．     

Characterization of a carbohydrate transporter from symbiotic glomeromycotan fungi

The symbiotic relationships between mycorrhizal fungi and plants have an enormous impact on terrestrial ecosystems. Most common are the arbuscular mycorrhizas, formed by fungi belonging to the phylum Glomeromycota. Arbuscular mycorrhizal fungi facilitate the uptake of soil nutrients by plants and in exchange obtain carbohydrates, thus representing a large sink for atmospheric plant-fixed CO(2). However, how carbohydrates are transported through the symbiotic interface is still unknown. Here we report the characterization of the first known glomeromycotan monosaccharide transporter, GpMST1, by exploiting the unique symbiosis of a glomeromycotan fungus (Geosiphon pyriformis) with cyanobacteria. The GpMST1 gene has a very low GC content and contains six introns with unusual boundaries. GpMST1 possesses twelve predicted transmembrane domains and functions as a proton co-transporter with highest affinity for glucose, then mannose, galactose and fructose. It belongs to an as yet uncharacterized phylogenetic monosaccharide transporter clade. This initial characterization of a new transporter family involved in fungal symbiosis will lead to a better understanding of carbon flows in terrestrial environments.     

benztropine类多巴胺转运蛋白配体的QSAR研究

利用Hansch方法，研究了37种benztropine类化合物diphenylmethoxy部位苯环取代基结构与其活性的定量关系，结果表明：苯环取代基的电性和体积等均是影响该类化合物与多巴胺转运蛋白亲和力的重要因素，所得到的benztropine类化合物diphenylmethoxy部位取代基的综合结构效应，对进一步研究该部位与多巴胺转运蛋白相互作用的电性和立体性质及设计新的多巴胺转运蛋白配体具有指导意义。     

MHC class I surface expression in embryo-derived cell lines inducible with peptide or interferon.

It has long been recognized that the absence of expression of products of the major histocompatibility complex (MHC) during early development might allow the fetus to escape recognition by maternal lymphocytes. In addition to the MHC class I heavy chain and beta 2-microglobulin, antigenic peptide is an essential structural component of the class I molecule. Indeed, there is evidence that MHC-linked genes encoding peptide transporter molecules and possibly components of a proteolytic complex are necessary for MHC class I assembly and stability at the cell surface. Here we demonstrate that embryonic cells in general show a defect in MHC class I assembly. Surface expression was rescued in the presence of an appropriate antigenic peptide, or by treatment with interferon. Consistent with this, HAM1 messenger RNA was not constitutively expressed, but was inducible by interferon, and during differentiation in vitro. Thus, tolerance of the fetal allograft may in part be controlled at the level of peptide-dependent MHC class I assembly.