解偶联蛋白4的线粒体保护作用

线粒体解偶联蛋白(UCPs)是近年来发现的线粒体膜蛋白家族中的新成员.研究表明,解偶联蛋白4(UCP4)有调节线粒体膜电位、减少氧自由基的生成、调节细胞内钙离子浓度等作用,受细胞代谢、甲状腺激素,以及儿茶酚胺等调节.UCP4主要分布于大脑皮质和海马区,可能在脑血管病、精神分裂症、变性病等线粒体易受损的疾病中起重要作用.     

瘦蛋白调控小型哺乳动物产热的分子机制

瘦蛋白(leptin)是由脂肪细胞分泌的、肥胖基因编码的蛋白类激素,其在哺乳动物的体重调节和适应性产热中有重要作用。解偶联蛋白(uncoupling protein,UCP)是动物体内具有解偶联作用的一类蛋白质。UCP-1是分布于褐色脂肪组织(brown adipose tissue,BAT)线粒体内膜上的标志蛋白,其在冷适应和食物资源缺乏等条件下的产热作用已得到印证。UCP-2和UCP-3的生物学功能仍存在争论。瘦蛋白可以通过诱导UCP的表达来促进产热．瘦蛋白诱导UCP表达的机制有二：(1)瘦蛋白通过与下丘脑神经肽Y的拮抗作用诱导UCP的表达;(2)瘦蛋白以交感神经系统为媒介诱导UCP的表达。瘦蛋白诱导UCP-1的表达是调节BAT产热的主要途径。了解瘦蛋白与UCP在产热中的作用对于我们了解动物产热的分子机制和其对低温、营养缺乏等条件下的生理生态适应机制具有重要意义。     

冷暴露对中缅树鼩褐色脂肪组织中解偶联蛋白1含量的影响

自备抗血清采用酶联免疫法测定了中缅树鼩(Tupaia belangeri)在(5±1)℃冷暴露0 d、7 d、14 d、21d、28 d时,褐色脂肪组织(BAT)中解偶联蛋白1(UCP1)的含量.结果表明,随着冷暴露时间的延长,中缅树鼩的体重、褐色脂肪组织重量均表现出了增加的趋势,BAT线粒体总蛋白和UCP1的含量也呈增加的趋势,其中UCP1的含量在28 d时达到极显著水平,比对照组增加了55.9%.说明冷暴露能够诱导中缅树鼩UCP1表达增加,从而使其适应性产热增加.     

“PM20D1”与“N酰基氨基酸”途径——新发现的呼吸链解偶联产热机制

正最近,一种新的呼吸链解偶联产热机制,被哈佛大学科学家公诸于世。经典呼吸链理论认为:解偶联蛋白-1(uncoupling protein 1,UCP1)是机体主要产热因子,其特异性表达在棕色脂肪细胞、米色脂肪细胞的线粒体内膜,并可将线粒体外膜H+势能转化为热能;由于该过程无ATP产生,而只产生热量,因此,亦称为解偶联呼吸(uncoupled respiration)。哈佛大学Dana-Farber癌症中心Spiegelman博士领导的研究组,首次发现     

解偶联蛋白2在心血管疾病发生发展中作用的研究现状

解偶联蛋白2(uncoupling protein 2,UCP2)是线粒体内膜质子载体蛋白,广泛存在多种组织和器官中。其通过降低线粒体内膜质子梯度,使呼吸作用中氧化磷酸化过程解偶联,从而发挥调控能量代谢、糖脂代谢和氧化应激等作用。近年来的研究发现,UCP2在心力衰竭、冠心病、高血压、肺动脉高压等疾病中也发挥重要作用。本文将对UCP2在心血管系统疾病发病中可能作用的研究现状作一综述。     

解偶联蛋白2对血管内皮的保护作用

解偶联蛋白2(UCP2)是核DNA编码的线粒体内膜阴离子转运体,广泛存在多种组织和器官中。其通过耗散线粒体内膜质子梯度发挥可诱导的解偶联作用。内皮细胞损伤是多种血管疾病的始动环节,近年来的研究发现,UCP2在动脉粥样硬化、高血压、糖尿病等中发挥血管内皮保护作用。本文对UCP2内皮保护作用的相关机制作一综述。     

解偶联蛋白与肥胖及2型糖尿病发病的关系

解偶联蛋白（UCPs）是线粒体内膜上的一种转运蛋白,它能够降低线粒体内膜上的质子梯度,使底物氧化和ADP磷酸化解偶联,减少ATP的产生。基于其功能,解偶联蛋白基因被视为肥胖病及2型糖尿病的重要候选基因。UCP同系物过表达的遗传工程小鼠表现出对饮食导致的肥胖具有耐受性,同时UCP2基因3＇非翻译区的45bp插入／缺失以及UCP3基因C-55T多态与肥胖表型的相关性等研究结论支持这一假说。本文对UCP基因与多基因控制的肥胖病及2型糖尿病发病的相关研究进行综述和讨论。 Abstract:Uncoupling proteins (UCPs) are mitochondria carrier proteins,which are able to dissipate the proton gradient of the inner mitochondria membrane.The uncoupling procedure reduces the amount of ATP generated through an oxidation of fuels.Therefore,UCPs are suggested as candidate genes for human obesity or type II diabetes mellitus.Experimental evidences,that genetically engineered mice over expressing different UCP homologues were resistant to diet-induced obesity and 45bp insertion polymorphism in the UCP2 3＇untranslated region and C-55T in UCP3 promoter region were associated with obesity related phenotype,supported the hypothesis.The roles of UCP genes in polygenic obesity and type II diabetes are evaluated and discussed in this paper.     

鲢鱼解偶联蛋白2全长cDNA序列的克隆及其组织表达

从淡水食毒藻鱼类鲢鱼(Hypophthalmichthys molitrix)肝脏，通过简并引物克隆解偶联蛋白2(uncoupling protein 2，UCP2) cDNA核心序列，应用5′RACE和3′RACE技术分别扩增该序列的5′末端和3′末端序列，最后通过序列拼接获得鲢鱼肝脏UCP2 cDNA全序列。序列分析结果表明，鲢鱼肝脏UCP2 cDNA全长1 452 bp，其中5′-UTR长337 bp，3′-UTR长182 bp，编码区933 bp，编码310个氨基酸，推测的氨基酸序列包含线粒体内膜载体蛋白3个特征结构及解偶联蛋白(UCPs)的特征序列。对鲢鱼不同组织UCP2的表达调控研究发现，鲢鱼组织UCP2基因在肠道、肝脏、肌肉、脂肪组织均大量表达，而在脑组织表达量较低，这与鲢鱼体内微囊藻毒素在这几个组织的分布完全一致，表明UCP2的功能可能与抑制微囊藻毒素引发过量活性氧(ROS)生成有关。     

解偶联蛋白2与衰老

解偶联蛋白(uncoupling protein,UCP)属于线粒体内膜上的一类载体蛋白,其生理作用是消除线粒体膜电位,使氧化磷酸化解偶联,从而抑制酸腺苷(adenosine triphosphate,ATP)合成,能量以热能形式散发.研究发现UCP2具有一种质子漏功能,表现对线粒体活性氧(reactive oxygen species,ROS)产生的调控和降低ROS的功能.在不同组织器官,不同代谢状态下UCP2的生理功能对细胞的影响不完全相同.特别是近年来的研究发现,UCP2参与了能量代谢、ROS的产生、子宫内膜退化、衰老等过程,并且与非酒精性脂肪肝、抗肥胖、动脉粥样硬化、局部缺血以及缺血再灌注损伤和2型糖尿病等有一定的相关性,倍受人们的关注.     

解偶联蛋白及功能研究进展

解偶联蛋白(ucP,uncoupling protein)是一类线粒体内膜上的载体,属于线粒体载体超家族,可以将H^ 从线粒体内膜渗漏到线粒体基质中,减少ATP的合成并产生热能。已知UCPl在小鼠中有维持体温和能量稳态的重要作用。而UCP2和UCP3可控制活性氧(reactive oxygen species,ROS)产生、调节脂肪酸氧化,并且在肥胖和糖尿病发生中有重要作用。     

Symmetric primary and tertiary structure mutations within a symmetric superfold: a solution, not a constraint, to achieve a foldable polypeptide

In previous studies designed to increase the primary structure symmetry within the hydrophobic core of human acidic fibroblast growth factor (FGF-1) a combination of five mutations were accommodated, resulting in structure, stability and folding kinetic properties similar to wild-type (despite the symmetric constraint upon the set of core residues). A sixth mutation in the core, involving a highly conserved Met residue at position 67, appeared intolerant to substitution. Structural analysis suggested that the local packing environment of position 67 involved two regions of apparent insertions that distorted the tertiary structure symmetry inherent in the beta-trefoil architecture. It was postulated that a symmetric constraint upon the primary structure within the core could only be achieved after these insertions had been deleted (concomitantly increasing the tertiary structure symmetry). The deletion of these insertions is now shown to permit mutation of position 67, thereby increasing the primary structure symmetry relationship within the core. Furthermore, despite the imposed symmetric constraint upon both the primary and tertiary structure, the resulting mutant form of FGF-1 is substantially more stable. The apparent inserted regions are shown to be associated with heparin-binding functionality; however, despite a marked reduction in heparin-binding affinity the mutant form of FGF-1 is surprisingly approximately 70 times more potent in 3T3 fibroblast mitogenic assays. The results support the hypothesis that primary structure symmetry within a symmetric protein superfold represents a possible solution, rather than a constraint, to achieving a foldable polypeptide.     

Integrating statistical pair potentials into protein complex prediction

The biophysical study of protein-protein interactions and docking has important implications in our understanding of most complex cellular signaling processes. Most computational approaches to protein docking involve a tradeoff between the level of detail incorporated into the model and computational power required to properly handle that level of detail. In this work, we seek to optimize that balance by showing that we can reduce the complexity of model representation and thus make the computation tractable with minimal loss of predictive performance. We also introduce a pair-wise statistical potential suitable for docking that builds on previous work and show that this potential can be incorporated into our fast fourier transform-based docking algorithm ZDOCK. We use the Protein Docking Benchmark to illustrate the improved performance of this potential compared with less detailed other scoring functions. Furthermore, we show that the new potential performs well on antibody-antigen complexes, with most predictions clustering around the Complementarity Determining Regions of antibodies without any manual intervention.     

The heat capacity of proteins

The heat capacity plays a major role in the determination of the energetics of protein folding and molecular recognition. As such, a better understanding of this thermodynamic parameter and its structural origin will provide new insights for the development of better molecular design strategies. In this paper we have analyzed the absolute heat capacity of proteins in different conformations. The results of these studies indicate that three major terms account for the absolute heat capacity of a protein: (1) one term that depends only on the primary or covalent structure of a protein and contains contributions from vibrational frequencies arising from the stretching and bending modes of each valence bond and internal rotations; (2) a term that contains the contributions of noncovalent interactions arising from secondary and tertiary structure; and (3) a term that contains the contributions of hydration. For a typical globular protein in solution the bulk of the heat capacity at 25°C is given by the covalent structure term (close to 85% of the total). The hydration term contributes about 15 and 40% to the total heat capacity of the native and unfolded states, respectively. The contribution of non-covalent structure to the total heat capacity of the native state is positive but very small and does not amount to more than 3% at 25°C. The change in heat capacity upon unfolding is primarily given by the increase in the hydration term (about 95%) and to a much lesser extent by the loss of noncovalent interactions (up to ～5%). It is demonstrated that a single universal mathematical function can be used to represent the partial molar heat capacity of the native and unfolded states of proteins in solution. This function can be experimentally written in terms of the molecular weight, the polar and apolar solvent accessible surface areas, and the total area buried from the solvent. This unique function accurately predicts the different magnitude and temperature dependences of the heat capacity of both the native and unfolded states, and therefore of the heat capacity changes associated with folding/unfolding transitions. © 1995 Wiley-Liss, Inc.     

Lethal mutations in the major homology region and their suppressors act by modulating the dimerization of the rous sarcoma virus capsid protein C‐terminal domain

An infective retrovirus requires a mature capsid shell around the viral replication complex. This shell is formed by about 1500 capsid protein monomers, organized into hexamer and pentamer rings that are linked to each other by the dimerization of the C‐terminal domain (CTD). The major homology region (MHR), the most highly conserved protein sequence across retroviral genomes, is part of the CTD. Several mutations in the MHR appear to block infectivity by preventing capsid formation. Suppressor mutations have been identified that are distant in sequence and structure from the MHR and restore capsid formation. The effects of two lethal and two suppressor mutations on the stability and function of the CTD were examined. No correlation with infectivity was found for the stability of the lethal mutations (D155Y‐CTD, F167Y‐CTD) and suppressor mutations (R185W‐CTD, I190V‐CTD). The stabilities of three double mutant proteins (D155Y/R185W‐CTD, F167Y/R185W‐CTD, and F167Y/I190V‐CTD) were additive. However, the dimerization affinity of the mutant proteins correlated strongly with biological function. The CTD proteins with lethal mutations did not dimerize, while those with suppressor mutations had greater dimerization affinity than WT‐CTD. The suppressor mutations were able to partially correct the dimerization defect caused by the lethal MHR mutations in double mutant proteins. Despite their dramatic effects on dimerization, none of these residues participate directly in the proposed dimerization interface in a mature capsid. These findings suggest that the conserved sequence of the MHR has critical roles in the conformation(s) of the CTD that are required for dimerization and correct capsid maturation. Proteins 2013. © 2012 Wiley Periodicals, Inc.     

A large scale test of computational protein design: folding and stability of nine completely redesigned globular proteins

A previously developed computer program for protein design, RosettaDesign, was used to predict low free energy sequences for nine naturally occurring protein backbones. RosettaDesign had no knowledge of the naturally occurring sequences and on average 65% of the residues in the designed sequences differ from wild-type. Synthetic genes for ten completely redesigned proteins were generated, and the proteins were expressed, purified, and then characterized using circular dichroism, chemical and temperature denaturation and NMR experiments. Although high-resolution structures have not yet been determined, eight of these proteins appear to be folded and their circular dichroism spectra are similar to those of their wild-type counterparts. Six of the proteins have stabilities equal to or up to 7kcal/mol greater than their wild-type counterparts, and four of the proteins have NMR spectra consistent with a well-packed, rigid structure. These encouraging results indicate that the computational protein design methods can, with significant reliability, identify amino acid sequences compatible with a target protein backbone.     

蛋白质相互作用研究的新技术与新方法

目前,蛋白质相互作用已成为蛋白质组学研究的热点. 新方法的建立及对已有技术的改进标志着蛋白质相互作用研究的不断发展和完善．在技术改进方面,本文介绍了弥补酵母双杂交的蛋白定位受限等缺陷的细菌双杂交系统;根据目标蛋白特性设计和修饰TAP标签来满足复合体研究要求的串联亲和纯化技术,以及在双分子荧光互补基础上发展的动态检测多个蛋白质间瞬时、弱相互作用的多分子荧光互补技术．还综述了近两年建立的新方法：与免疫共沉淀相比,寡沉淀技术直接研究具有活性的蛋白质复合体;减量式定量免疫沉淀方法排除了蛋白质复合体中非特异性相互作用的干扰;原位操作的多表位－配基绘图法避免了样品间差异的影响,以及利用多点吸附和交联加固研究弱蛋白质相互作用的固相蛋白质组学方法.     

Human dihydrofolate reductase and thymidylate synthase form a complex in vitro and co-localize in normal and cancer cells

Enzymes involved in thymidylate biosynthesis, thymidylate synthase (TS), and dihydrofolate reductase (DHFR) are well-known targets in cancer chemotherapy. In this study, we demonstrated for the first time, that human TS and DHFR form a strong complex in vitro and co-localize in human normal and colon cancer cell cytoplasm and nucleus. Treatment of cancer cells with methotrexate or 5-fluorouracil did not affect the distribution of either enzyme within the cells. However, 5-FU, but not MTX, lowered the presence of DHFR-TS complex in the nucleus by 2.5-fold. The results may suggest the sequestering of TS by FdUMP in the cytoplasm and thereby affecting the translocation of DHFR-TS complex to the nucleus. Providing a strong likelihood of DHFR-TS complex formation in vivo, the latter complex is a potential new drug target in cancer therapy. In this paper, known 3D structures of human TS and human DHFR, and some protozoan bifunctional DHFR-TS structures as templates, are used to build an in silico model of human DHFR–TS complex structure, consisting of one TS dimer and two DHFR monomers. This complex structure may serve as an initial 3D drug target model for prospective inhibitors targeting interfaces between the DHFR and TS enzymes.     

Characterization of a mammalian Golgi-localized protein complex, COG, that is required for normal Golgi morphology and function

Multiprotein complexes are key determinants of Golgi apparatus structure and its capacity for intracellular transport and glycoprotein modification. Three complexes that have previously been partially characterized include (a) the Golgi transport complex (GTC), identified in an in vitro membrane transport assay, (b) the ldlCp complex, identified in analyses of CHO cell mutants with defects in Golgi-associated glycosylation reactions, and (c) the mammalian Sec34 complex, identified by homology to yeast Sec34p, implicated in vesicular transport. We show that these three complexes are identical and rename them the conserved oligomeric Golgi (COG) complex. The COG complex comprises four previously characterized proteins (Cog1/ldlBp, Cog2/ldlCp, Cog3/Sec34, and Cog5/GTC-90), three homologues of yeast Sec34/35 complex subunits (Cog4, -6, and -8), and a previously unidentified Golgi-associated protein (Cog7). EM of ldlB and ldlC mutants established that COG is required for normal Golgi morphology. "Deep etch" EM of purified COG revealed an approximately 37-nm-long structure comprised of two similarly sized globular domains connected by smaller extensions. Consideration of biochemical and genetic data for mammalian COG and its yeast homologue suggests a model for the subunit distribution within this complex, which plays critical roles in Golgi structure and function.     

A review of visualisations of protein fold networks and their relationship with sequence and function

Proteins form arguably the most significant link between genotype and phenotype. Understanding the relationship between protein sequence and structure, and applying this knowledge to predict function, is difficult. One way to investigate these relationships is by considering the space of protein folds and how one might move from fold to fold through similarity, or potential evolutionary relationships. The many individual characterisations of fold space presented in the literature can tell us a lot about how well the current Protein Data Bank represents protein fold space, how convergence and divergence may affect protein evolution, how proteins affect the whole of which they are part, and how proteins themselves function. A synthesis of these different approaches and viewpoints seems the most likely way to further our knowledge of protein structure evolution and thus, facilitate improved protein structure design and prediction.     

Improvement of protein stability in protein microarrays

Protein stability in microarrays was improved using protein stabilizers. PEG 200 at 30% (w/v) was the most efficient stabilizer giving over 4-fold improvement in protein stability compared to without the stabilizer. PEG 200 above 10% (w/v) in the array solution prevented the evaporation of water in the sample and thereby improved protein stability in the microarray. When the streptavidin-biotin binding reaction was performed under optimized conditions, biotin-BSA-fluorescein isothiocyanate (FITC) was detected from 1 ng ml^–1 to 5 g ml^–1 by fluorescence analysis.