基于序列剖面和可及表面积的蛋白质相互作用位点的预测

蛋白质相互作用位点的预测对于突变设计和蛋白质相互作用网络的重构都是至关重要的.由于实验确定的蛋白质复合物和蛋白质配体复合物的结构依然相当少,预测蛋白质相互作用位点的计算方法就显得十分重要.该文提出了一种以支持向量机为分类器,以邻近残基的序列剖面和可及表面积为输入数据来预测蛋白质相互作用位点的方法.计算结果显示,界面残基和非界面残基被识别的准确率为75.12%,假阳性率为28.04%.与输入数据仅有序列剖面的方法相比,界面残基和非界面残基被识别的准确率提高了4.34%,假阳性率降低了4.63%.     

四川地区利福平耐药结核分枝杆菌rpoB基因利福平耐药决定区突变分析

本文对268株结核分枝杆菌临床分离株(包括213株利福平耐药株、55株利福平敏感株)rpoB基因RRDR区进行测序并比较分析.结果表明,在213株利福平耐药株中有200株(93.90%)在rpoB基因RRDR区发生了突变,在55株利福平敏感株中RRDR均未发生突变.突变频率最高的类型依次为:531位点(55.87%),526位点(16.43%),516位点(10.33%)和511位点(8.92%),以上四个位点的突变菌株数占所有利福平耐药株的91.55%.四川地区利福平耐药株中511位点突变频率高于全国其他地区.     

基于预训练蛋白质语言模型的氨基酸致病突变预测

依赖于临床标签的氨基酸致病突变预测方法通常由于标签存在跨基因的偏差、稀疏噪声等因素，出现性能膨胀的情况.为解决此问题，创新地在不需要标签的情况下，利用预训练蛋白质语言模型计算ClinVar数据库中突变位点的氨基酸概率分布，并基于此分布构造突变型与野生型氨基酸出现概率的对数优势比（LOR），使用一种全局-局部结合的高斯混合模型拟合LOR，从而无监督地计算突变致病效应概率分数（PPE）并推断致病性，最后给出预测的不确定性度量.使用与深度突变扫描（DMS）实验的相关性作为评估指标以避免标签泄漏等问题.模型评估结果验证PPE具有稳健的致病性预测性能，在2458个蛋白质上的接收者操作特征曲线下面积（AUC）平均值约为0.89，与4种DMS实验的平均斯皮尔曼相关系数约为0.44，优于大部分依赖标签的计算方法，且与高通量实验的性能相当.该研究为遗传变异的解释、疾病的研究、诊断和临床治疗提供了可靠的辅助工具.     

同源建模方法预测蛋白质突变结构的适用性分析

通过从Protein Data Bank(PDB)结构数据库中提取单氨基酸突变的晶体结构,构建了一组无冗余的测试数据集,对目前应用最广泛的两款同源建模预测软件(SWISS-MODEL和MODELLER)进行了测试分析,发现它们对蛋白质的整体结构预测效果良好,均方根偏差小于0.5埃(RMSD0.5),但在突变导致结构显著变化(RMSD1.5)的情况下却均不能得到准确结果.分类统计显示,发生在蛋白质结构内部和极性氨基酸之间的突变结构变化小,两款软件预测效果较好(RMSD1.0).突变导致结构显著变化的可能性不高(5%),但它对蛋白质功能的影响不可忽视,因此应用同源建模方法对于蛋白质突变的模拟并不完全适用,还需要开发新方法来提高准确性.     

人类Batten病基因CLN3及其突变的生物信息学分析

利用生物信息学方法对与儿童蜡样质脂褐质沉积症(NCLs)相关基因CLN3的结构进行了深入的分析.根据CLN3的Mrna序列对其进行了染色体精确定位,确定CLN3的ORF及其外显子的位置;分析CLN3蛋白质(CLN 3P)结构特点和修饰位点,并明确近缘种及远缘种的保守区域和突变致病的关系.结果表明,CLN3位于人类16号染色体的短臂(16(p12.1～p11.2));基因全长为14804bp,包含15个外显子和14个内含子,ORF长度1317bp,编码438个氨基酸.蛋白质功能域分析表明,CLN 3P是一种分子量为43kDa的高度糖基化的溶酶体膜蛋白,该蛋白存在酰基化、糖基化和多种磷酸化修饰位点,在近缘的CLN 3P中存在高度保守的区域,进一步对10个远缘蛋白的多序列进行比对,发现了该蛋白质中最为保守的区域;45种CLN 3P的突变分析显示,主要的突变位于101、161～162、170、187、199、211、295、327、330、334、352等12个氨基酸位点,这些突变几乎发生在保守区域,突变还表现出一定的分布特点及地区差异,说明保守序列的突变是该疾病产生的重要原因,为该病的分子诊断提供了依据.     

云南妇女卵巢癌中p21WAF基因有较高的31位密码(Ser/Arg)突变

 通过PCR-SSCP分析和序列分析,对取自云南地区的44例妇女卵巢肿瘤样本的p21^WAF基因Ser³¹密码子的突变进行了分析.结果表明35例恶性肿瘤样本中有15例发现有突变,突变率为42.9%;9例良性肿瘤中有4例出现突变,突变频率44.4%;在另外6例正常组织中出现1例突变,突变率为16.7%.这一结果表明,云南妇女在这一基因位点存在多态性,这一位点的突变具有发生卵巢癌的趋势.     

基于突变理论的图像边缘检测技术

边缘检测是图像处理的基本问题之一,它为人们描述或识别目标以及解释图像提供了一个重要的特征参数.利用突变理论研究图像的突变特性,利用突变模型描述系统状态变化的规律,通过尖点突变理论建立边缘模型,来实现有效的图像边缘检测.实验结果表明,提出的边缘检测方法,能够准确检测到连续的图像边缘且具有较好的实时性.     

在线传感器突变信号的检测与区分

为了准确区分传感器突变信号产生的原因,提出了基于数学模型的小波频带分析法. 针对工业流程中的测控系统,分析了输出突变信号的频率组成与突变原因的关系. 用小波频带分析技术,将高低频信号分离,并进行能量统计,根据高低频信号能量比例的变化,判断出突变信号产生的原因. 经典型控制系统的计算机仿真和恒压供水系统实验结果表明,该方法能够有效地诊断出传感器是否发生故障.     

甘露(聚)糖结合凝集素外显子Ⅰ 54位密码子突变的检测方法

对人源甘露糖结合凝集素基因54位密码子点突变的情况进行测定,建立了相关实验室检测方法.采用PCR扩增目的片段,再对产物采用PCR-RFLP测定点突变.检测的哈萨克40例血样54位密码子的点突变情况:野生型为20例,占50%;突变杂合型为20,占50%;突变纯合子为0例,占0%.PCR-RFLP适合实验室检测MBL外显子I54密码子突变情况,可以作为分析依据.     

中国内蒙古通辽地区和吉林长春地区宫颈癌中HPV16型E6基因变异分析

为了解我国内蒙古通辽和吉林长春地区宫颈癌中HPV16亚型的分布情况,对通辽和长春的30例HPV16阳性宫颈癌样品中的HPV16 E6基因进行序列分析.结果在30例宫颈癌样品中的E6全长基因内发现11处位点突变,形成了14种突变体,其中突变T459G是本试验第一次发现的突变.经系统进化分析发现,所有突变体均属欧洲原型突变群.结合之前我国HPV16亚型分布调查结果表明,欧洲型和东亚型HPV16在我国不同地区,分布不均一,主要以欧洲原型突变群为主.     

Differential expression of calmodulin-binding proteins in B, T lymphocytes and thymocytes

Changes in intracellular free Ca2+ are involved in the transmembrane signalling of different cells, including lymphocytes. Since calmodulin (CaM) is a primary receptor for Ca2+ (ref. 4), it may mediate the activation of crucial enzymes after antigen-induced increases in cytosolic Ca2+. Using a biotinylated-CaM (Bio-CaM) detection procedure to identify such proteins, we found that a peptide of relative molecular mass 59,000 (59K) was the predominant soluble CaM-binding protein (CaM-BP) in T cells and B lymphocytes from murine spleen; immunoblotting experiments identified it as a subunit of the CaM-dependent phosphatase, 'calcineurin' (CN). Smaller amounts of larger CaM-BPs, thought to be cytoskeletal-binding proteins, were also detected. CaM-BPs were expressed differentially, with B lymphocytes having four times more of the CN-like protein than T lymphocytes, while in thymocytes, a 65K polypeptide was the major CaM-BP. However, limited proteolysis analysis suggested that this thymus-specific peptide may be a precursor of CN. These data suggest that Ca2+-stimulated protein dephosphorylation may be an important and highly regulated function in lymphoid cells.     

Mutations of the BRAF gene in human cancer

Cancers arise owing to the accumulation of mutations in critical genes that alter normal programmes of cell proliferation, differentiation and death. As the first stage of a systematic genome-wide screen for these genes, we have prioritized for analysis signalling pathways in which at least one gene is mutated in human cancer. The RAS RAF MEK ERK MAP kinase pathway mediates cellular responses to growth signals. RAS is mutated to an oncogenic form in about 15% of human cancer. The three RAF genes code for cytoplasmic serine/threonine kinases that are regulated by binding RAS. Here we report BRAF somatic missense mutations in 66% of malignant melanomas and at lower frequency in a wide range of human cancers. All mutations are within the kinase domain, with a single substitution (V599E) accounting for 80%. Mutated BRAF proteins have elevated kinase activity and are transforming in NIH3T3 cells. Furthermore, RAS function is not required for the growth of cancer cell lines with the V599E mutation. As BRAF is a serine/threonine kinase that is commonly activated by somatic point mutation in human cancer, it may provide new therapeutic opportunities in malignant melanoma.     

Structural Prediction of Membrane Protein: Application to Known Structures

Membrane proteins are embedded in the lipid bilayer,which creates a suitable environment for their actions. It is important to decide which tpye it belongs to because it is closely relevant to its biological fumction and its interaction process with other molecules in a biological system. Membrane proteins have different types. The function of a membrane protein is closely correlated with the type it belongs to. In this study, on the basis of the concept of pseudo amino acid (PseAA) composition originally introduced by Chou, the value of approximate entropy (ApEn) of the query membrane protein was used to integrate the complementary information. By fusing fifteen powerful individual fuzzy K-nearest neighbor (FKNN) classifiers, an ensemble classifier was presented.Each basic classifier was trained in PseAA composition of membrane protein sequences with different parameters. The results of experiments demonstrate it is efficient for the structural prediction of membrane proteins.     

原发性子宫内膜癌与p16基因突变及蛋白表达关系的研究

目的研究p16蛋白表达与原发性子宫内膜癌发生发展的关系和p16基因缺失突变及点突变在原发性子宫内膜癌发生发展中的地位.方法利用免疫组织化学方法、聚合酶链反应和聚合酶链反应-单链构象多态性分析技术,分别检测正常子宫内膜组织、子宫内膜癌前病变组织及原发性子宫内膜癌组织,观察p16蛋白和p16基因缺失突变及点突变.结果1)p16蛋白阳性表达率在正常子宫内膜组织和子宫内膜癌前病变组织中表达率分别为92.78%和90.00%,两者相比无差异性;在原发性子宫内膜癌组织中为66.67%,明显低于正常子宫组织及癌前病变组织;2)在42例原发性性子宫内膜癌组织中有14例发生p16基因缺失突变,4例发生了p16基因点突变,突变率为分别为33.33%和9.6%,正常子宫颈组织和子宫内膜癌前病变组织未发现p16基因缺失突变和点突变.结论1)在原发性子宫内膜癌发生发展过程中,p16蛋白的表达在高分化癌明显低于低分化癌,表明p16蛋白缺乏与子宫颈细胞增殖失控及分化不良紧密相关.2)原发性子宫内膜癌存在p16基因点突变,以低分化癌多见,但不是较频繁的事件;原发性子宫内膜癌存在p16基因缺失突变,以低分化癌多见,是较频繁的事件.3)p16蛋白表达与p16基因突变的相关性未被发现.     

F-box proteins in flowering plants

In eukaryotes, the ubiquitin-mediated protein degradation pathway has been shown to control several key biological processes such as cell division, development, metabolism and immune response. F-box proteins, as a part of SCF (Skp1-Cullin (or Cdc53)-F-box) complex, functioned by interacting with substrate proteins, leading to their subsequent degradation by the 26S proteasome. To date, several F-box proteins identified in Arabidopsis and Antirrhinum have been shown to play important roles in auxin signal transduction, floral organ formation, flowering and leaf senescence. Arabidopsis genome sequence analysis revealed that it encodes over 1000 predicted F-box proteins accounting for about 5% of total predicted proteins. These results indicate that the ubiquitin-mediated protein degradation involving the F-box proteins is an important mechanism controlling plant gene expression. Here, we review the known F-box proteins and their functionsin flowering plants.     

Evolution of increased complexity in a molecular machine

Many cellular processes are carried out by molecular 'machines'-assemblies of multiple differentiated proteins that physically interact to execute biological functions. Despite much speculation, strong evidence of the mechanisms by which these assemblies evolved is lacking. Here we use ancestral gene resurrection and manipulative genetic experiments to determine how the complexity of an essential molecular machine--the hexameric transmembrane ring of the eukaryotic V-ATPase proton pump--increased hundreds of millions of years ago. We show that the ring of Fungi, which is composed of three paralogous proteins, evolved from a more ancient two-paralogue complex because of a gene duplication that was followed by loss in each daughter copy of specific interfaces by which it interacts with other ring proteins. These losses were complementary, so both copies became obligate components with restricted spatial roles in the complex. Reintroducing a single historical mutation from each paralogue lineage into the resurrected ancestral proteins is sufficient to recapitulate their asymmetric degeneration and trigger the requirement for the more elaborate three-component ring. Our experiments show that increased complexity in an essential molecular machine evolved because of simple, high-probability evolutionary processes, without the apparent evolution of novel functions. They point to a plausible mechanism for the evolution of complexity in other multi-paralogue protein complexes.     

天人菊matK基因克隆及生物信息学分析

为了完善天人菊的遗传信息,丰富菊科植物分子生物学分析数据.通过PCR扩增、基因克隆获得天人菊matK基因的完整核苷酸序列,采用生物信息学方法分析天人菊matK蛋白的结构及性质,并与其他12种matK氨基酸序列进行对比,构建系统进化树.结果表明,天人菊matK基因全长1296 bp,可编码432个氨基酸,二级结构以α-螺...     

阿特拉津氯水解酶基因的定点诱变和酶活力检测

阿特拉津氯水解酶(AtzA)是一种对除草剂的生物降解和环境净化有重要意义的酶．采用定点诱变方法将假单胞菌ADP株的atzA基因第832位碱基(鸟嘌呤)诱变成腺嘌吟，然后将其插入表达载体pET21b( )，并在大肠杆菌中表达．表达的蛋白特性研究表明：AtzA或AtzA—NK融合蛋白系水溶性蛋白，很容易通过Ni—NTA Magnetic Agarose Beads分离纯化．采用阿特拉津脱氯反应产生HCl而引起pH指示剂颜色改变的测定方法能方便地对其酶活力进行定量．酶活力结果表明，突变酶的比活力与假单胞菌ADP菌株的AtzA相比没有明显改变，暗示突变位点(第278位缬氨酸突变成甲硫氨酸)不是酶的活性中心或底物结合部位。     

初渣流动方式对高炉内硅传递过程的影响

从初渣偏斜流动的观点出发,对高炉滴落带生铁中硅含量的变化过程进行了热力学分析和实验研究,探讨了初渣偏流与炉内硅还原过程的相互关系。结果表明,高炉内生铁中硅含量的变化规律受初渣偏流和高温下焦炭对 SiO_(g)的还原反应所控制,从而提出了合理的高炉冶炼低硅生铁的技术措施。     

Point mutation of an FGF receptor abolishes phosphatidylinositol turnover and Ca2+ flux but not mitogenesis.

Stimulation of certain receptor tyrosine kinases results in the tyrosine phosphorylation and activation of phospholipase C gamma (PLC gamma), an enzyme that catalyses the hydrolysis of phosphatidylinositol (PtdIns). This hydrolysis generates diacylglycerol and free inositol phosphate, which in turn activate protein kinase C and increase intracellular Ca2+, respectively. PLC gamma physically associates with activated receptor tyrosine kinases, suggesting that it is a substrate for direct phosphorylation by these kinases. Here we report that a fibroblast growth factor (FGF) receptor with a single point mutation at residue 766 replacing tyrosine with phenylalanine fails to associate with PLC gamma in response to FGF. This mutant receptor also failed to mediate PtdIns hydrolysis and Ca2+ mobilization after FGF stimulation. However, the mutant receptor phosphorylated itself and several other cellular proteins, and it mediated mitogenesis in response to FGF. These findings show that a point mutation in the FGF receptor selectively eliminates activation of PLC gamma and that neither Ca2+ mobilization nor PtdIns hydrolysis are required for FGF-induced mitogenesis.