Structural Variations in Protein Superfamilies: Actin and Tubulin

Structures of homologous proteins are usually conserved during evolution, as are critical active site residues. This is the case for actin and tubulin, the two most important cytoskeleton proteins in eukaryotes. Actins and their related proteins (Arps) constitute a large superfamily whereas the tubulin family has fewer members. Unaligned sequences of these two protein families were analysed by searching for short groups of family-specific amino acid residues, that we call motifs, and by counting the number of residues from one motif to the next. For each sequence, the set of motif-to-motif residue counts forms a subfamily-specific pattern (landmark pattern) allowing actin and tubulin superfamily members to be identified and sorted into subfamilies. The differences between patterns of individual subfamilies are due to inserts and deletions (indels). Inserts appear to have arisen at an early stage in eukaryote evolution as suggested by the small but consistent kingdom-dependent differences found within many Arp subfamilies and in γ-tubulins. Inserts tend to be in surface loops where they can influence subfamily-specific function without disturbing the core structure of the protein. The relatively few indels found for tubulins have similar positions to established results, whereas we find many previously unreported indel positions and lengths for the metazoan Arps.     

视黄醇结合蛋白的结构与功能

视黄醇结合蛋白（RBP）是视黄醇转运的载体蛋白,作为结合小分子流水物质的载体蛋白家族（lipocalin）的一个重要成员,其结构与功能的研究正受到国外学者的重视,文章介绍了视黄醇结合蛋白的性质、结构研究进展,讨论了视黄醇结合蛋白与前蛋白和受体相互作用的位点和结构特点．     

A Novel Method to Predict Genomic Islands Based on Mean Shift Clustering Algorithm

Genomic Islands (GIs) are regions of bacterial genomes that are acquired from other organisms by the phenomenon of horizontal transfer. These regions are often responsible for many important acquired adaptations of the bacteria, with great impact on their evolution and behavior. Nevertheless, these adaptations are usually associated with pathogenicity, antibiotic resistance, degradation and metabolism. Identification of such regions is of medical and industrial interest. For this reason, different approaches for genomic islands prediction have been proposed. However, none of them are capable of predicting precisely the complete repertory of GIs in a genome. The difficulties arise due to the changes in performance of different algorithms in the face of the variety of nucleotide distribution in different species. In this paper, we present a novel method to predict GIs that is built upon mean shift clustering algorithm. It does not require any information regarding the number of clusters, and the bandwidth parameter is automatically calculated based on a heuristic approach. The method was implemented in a new user-friendly tool named MSGIP—Mean Shift Genomic Island Predictor. Genomes of bacteria with GIs discussed in other papers were used to evaluate the proposed method. The application of this tool revealed the same GIs predicted by other methods and also different novel unpredicted islands. A detailed investigation of the different features related to typical GI elements inserted in these new regions confirmed its effectiveness. Stand-alone and user-friendly versions for this new methodology are available at http://msgip.integrativebioinformatics.me.     

Crystal Structure of Vaccinia Viral A27 Protein Reveals a Novel Structure Critical for Its Function and Complex Formation with A26 Protein

Vaccinia virus envelope protein A27 has multiple functions and is conserved in the Orthopoxvirus genus of the poxvirus family. A27 protein binds to cell surface heparan sulfate, provides an anchor for A26 protein packaging into mature virions, and is essential for egress of mature virus (MV) from infected cells. Here, we crystallized and determined the structure of a truncated form of A27 containing amino acids 21–84, C71/72A (tA27) at 2.2 Å resolution. tA27 protein uses the N-terminal region interface (NTR) to form an unexpected trimeric assembly as the basic unit, which contains two parallel α-helices and one unusual antiparallel α-helix; in a serpentine way, two trimers stack with each other to form a hexamer using the C-terminal region interface (CTR). Recombinant tA27 protein forms oligomers in a concentration-dependent manner in vitro in gel filtration. Analytical ultracentrifugation and multi-angle light scattering revealed that tA27 dimerized in solution and that Leu47, Leu51, and Leu54 at the NTR and Ile68, Asn75, and Leu82 at the CTR are responsible for tA27 self-assembly in vitro. Finally, we constructed recombinant vaccinia viruses expressing full length mutant A27 protein defective in either NTR, CTR, or both interactions; the results demonstrated that wild type A27 dimer/trimer formation was impaired in NTR and CTR mutant viruses, resulting in small plaques that are defective in MV egress. Furthermore, the ability of A27 protein to form disulfide-linked protein complexes with A26 protein was partially or completely interrupted by NTR and CTR mutations, resulting in mature virion progeny with increased plasma membrane fusion activity upon cell entry. Together, these results demonstrate that A27 protein trimer structure is critical for MV egress and membrane fusion modulation. Because A27 is a neutralizing target, structural information will aid the development of inhibitors to block A27 self-assembly or complex formation against vaccinia virus infection.     

Using Pair-Coupled Amino Acid Composition to Predict Protein Secondary Structure Content

The pair-coupled amino acid composition is introduced to predict the secondary structure contents of a protein. Compared with the existing methods all based on singlewise amino acid composition as defined in a 20D (dimensional) space, this represents a step forward to the consideration of the sequence coupling effect. The test results indicate that the introduction of the pair-coupled amino acid composition can significantly improve the prediction quality. It is anticipated that the concept of the pair-coupled amino acid composition can be used to simplify the formulation of sequence coupling (or sequence order) effects and to study many other features of proteins as well.     

蛋白质结构与功能中的结构域

结构域是蛋白质亚基结构中的紧密球状区域.结构域作为蛋白质结构中介于二级与三级结构之间的又一结构层次,在蛋白质中起着独立的结构单位、功能单位与折叠单位的作用.在复杂蛋白质中,结构域具有结构与功能组件与遗传单位的作用.结构域层次的研究将会促进蛋白质结构与功能关系、蛋白质折叠机制以及蛋白质设计的研究.     

A Novel Approach to Phylogeny Reconstruction from Protein Sequences

The reliable reconstruction of tree topology from a set of homologous sequences is one of the main goals in the study of molecular evolution. If consistent estimators of distances from a multiple sequence alignment are known, the distance method is attractive because the tree reconstruction is consistent. To obtain a distance estimate d, the observed proportion of differences p (p-distance) is usually ``corrected' for multiple and back substitutions by means of a functional relationship d=f(p). In this paper the conditions under which this correction of p-distances will not alter the selection of the tree topology are specified. When these conditions are not fulfilled the selection of the tree topology may depend on the correction function applied. A novel method which includes estimates of distances not only between sequence pairs, but between triplets, quadruplets, etc., is proposed to strengthen the proper selection of correction function and tree topology. A ``super' tree that includes all tree topologies as special cases is introduced. Received: 17 February 1998 / Accepted: 20 July 1998     

QUAFIT: A Novel Method for the Quaternary Structure Determination from Small-Angle Scattering Data

The new QUAFIT method for determining the quaternary structure of biological macromolecular assemblies by analyzing x-ray or neutron small-angle scattering data is presented. The method is based on the idea that asymmetric monomers, formed by rigid domains of known atomic structure possibly connected by flexible linkers of known sequence, are assembled according to a point-group symmetry combined with a screw axis. Scattering amplitudes of domains and linkers are determined by means of a spherical harmonics expansion and combined to get the form factor of the assembly. To avoid any overlap among domains, the contact distance between two asymmetric domains is determined as a function of their orientation by a new algorithm, based on Stone's Invariants expansion. To account for continuity and compactness of the whole assembly, an anisotropic Lennard-Jones potential among domains, written in terms of the contact distances, is included in the merit function. QUAFIT allows for the simultaneous presence of oligomerization intermediates as well as of monomers distributed over multiple conformations. QUAFIT has been tested by studying the structure of a high molecular weight protein, the hemocyanin from Octopus vulgaris, under solution conditions that stabilize the decameric form or induce dissociation into monomers, respectively. Results are in very good agreement with the structural model derived from electron microscopy observations.     

Interactome Mapping: Using Protein Microarray Technology to Reconstruct Diverse Protein Networks

A major focus of systems biology is to characterize interactions between cellular components, in order to develop an accurate picture of the intricate networks within biological systems. Over the past decade, protein microarrays have greatly contributed to advances in proteomics and are becoming an important platform for systems biology. Protein microarrays are highly flexible, ranging from large-scale proteome microarrays to smaller customizable microarrays, making the technology amenable for detection of a broad spectrum of biochemical properties of proteins. In this article, we will focus on the numerous studies that have utilized protein microarrays to reconstruct biological networks including protein-DNA interactions, posttranslational protein modifications (PTMs), lectin-glycan recognition, pathogen-host interactions and hierarchical signaling cascades. The diversity in applications allows for integration of interaction data from numerous molecular classes and cellular states, providing insight into the structure of complex biological systems. We will also discuss emerging applications and future directions of protein microarray technology in the global frontier.     

RRM RNA结合蛋白的结构与功能

RRM RNA结合蛋白是一类含一个或数个RRM结构域及附属结构域的RNA结合蛋白,参与RNA前体的剪接、RNA的细胞定位、RNA的稳定性等多种转录后调控过程.在RRM基序中含有许多保守的氨基酸以保证对RNA的结合活性,但是这一家族的不同蛋白质却能特异地结合各种不同的RNA分子.RRM RNA结合蛋白与某些人类遗传性疾病及肿瘤相关.     

Joint Evolutionary Trees: A Large-Scale Method To Predict Protein Interfaces Based on Sequence Sampling

The Joint Evolutionary Trees (JET) method detects protein interfaces, the core residues involved in the folding process, and residues susceptible to site-directed mutagenesis and relevant to molecular recognition. The approach, based on the Evolutionary Trace (ET) method, introduces a novel way to treat evolutionary information. Families of homologous sequences are analyzed through a Gibbs-like sampling of distance trees to reduce effects of erroneous multiple alignment and impacts of weakly homologous sequences on distance tree construction. The sampling method makes sequence analysis more sensitive to functional and structural importance of individual residues by avoiding effects of the overrepresentation of highly homologous sequences and improves computational efficiency. A carefully designed clustering method is parametrized on the target structure to detect and extend patches on protein surfaces into predicted interaction sites. Clustering takes into account residues' physical-chemical properties as well as conservation. Large-scale application of JET requires the system to be adjustable for different datasets and to guarantee predictions even if the signal is low. Flexibility was achieved by a careful treatment of the number of retrieved sequences, the amino acid distance between sequences, and the selective thresholds for cluster identification. An iterative version of JET (iJET) that guarantees finding the most likely interface residues is proposed as the appropriate tool for large-scale predictions. Tests are carried out on the Huang database of 62 heterodimer, homodimer, and transient complexes and on 265 interfaces belonging to signal transduction proteins, enzymes, inhibitors, antibodies, antigens, and others. A specific set of proteins chosen for their special functional and structural properties illustrate JET behavior on a large variety of interactions covering proteins, ligands, DNA, and RNA. JET is compared at a large scale to ET and to Consurf, Rate4Site, siteFiNDER|3D, and SCORECONS on specific structures. A significant improvement in performance and computational efficiency is shown.     

蛋白磷酸酶2A的结构、功能和活性调节

蛋白磷酸酶 2A(proteinphosphatase 2A ,PP2A)是主要的丝 /苏氨酸蛋白磷酸酶 ,拥有众多不同基因编码的亚基 ,分别组成多种不同的PP2A全酶 ,参与细胞周期、DNA复制、信号转导、细胞分化和细胞恶性转化等多种细胞生物学事件 ,并和神经退行性疾病、肿瘤等多种疾病的发生、发展有关。PP2A调节亚基的组织特异性表达和细胞内定位 ,催化亚基羧基末端的磷酸化和甲基化 ,第二信使神经酰胺 (ceramide)、天然小分子抑制剂等都能够调节PP2A的活性。     

2019新型冠状病毒S蛋白的结构和功能分析

运用生物信息学,预测急性呼吸系统综合症冠状病毒2(SARS-CoV-2/2019-nCoV)的基本理化性质、结构、功能和抗原表位等,为新型冠状病毒肺炎(COVID-19)的防治提供思路。应用ExPASy分析S蛋白的消光系数、不稳定系数和半衰期等理化性质;利用SignaIP v5.0分析S蛋白的信号肽;应用TMHMM分析S蛋白的跨膜区;利用NetPhos3.1在线工具预测S蛋白的磷酸化位点;应用Pfam预测S蛋白的结构域;应用PSIPRED分析S蛋白的二级结构特征;利用SWISS-MODEL构建S蛋白的三级结构;利用BLAST分析SARS-CoV-2的S蛋白与其他物种的相似性;利用MEGA软件分析2019-nCoV的S蛋白与其他物种的进化关系。S蛋白由1 273个氨基酸组成,其相对分子质量为141 178.47,等电点为6.24,含有一个跨膜区,是低亲水性分泌蛋白;S蛋白的基本组成单位为纤突蛋白,其二级结构中以无规则卷曲和螺旋结构为主,三级结构中纤突糖蛋白和ACE2复合体具有重要的意义;2019-nCoV与蝙蝠冠状病毒和SARS-CoV同源;S蛋白存在多个潜在的线性T细胞和B细胞表位,1 202～1 210位氨基酸区域的抗原性和应答频率最高。生物信息学技术有利于了解S蛋白的理化性质、结构、功能和潜在的线性T细胞表位等,可为新型冠状肺炎的研究和防治提供参考依据。     

A Novel Bioinformatics Strategy for Function Prediction of Poorly-Characterized Protein Genes Obtained from Metagenome Analyses

As a result of remarkable progresses of DNA sequencing technology, vast quantities of genomic sequences have been decoded. Homology search for amino acid sequences, such as BLAST, has become a basic tool for assigning functions of genes/proteins when genomic sequences are decoded. Although the homology search has clearly been a powerful and irreplaceable method, the functions of only 50% or fewer of genes can be predicted when a novel genome is decoded. A prediction method independent of the homology search is urgently needed. By analyzing oligonucleotide compositions in genomic sequences, we previously developed a modified Self-Organizing Map ‘BLSOM’ that clustered genomic fragments according to phylotype with no advance knowledge of phylotype. Using BLSOM for di-, tri- and tetrapeptide compositions, we developed a system to enable separation (self-organization) of proteins by function. Analyzing oligopeptide frequencies in proteins previously classified into COGs (clusters of orthologous groups of proteins), BLSOMs could faithfully reproduce the COG classifications. This indicated that proteins, whose functions are unknown because of lack of significant sequence similarity with function-known proteins, can be related to function-known proteins based on similarity in oligopeptide composition. BLSOM was applied to predict functions of vast quantities of proteins derived from mixed genomes in environmental samples.     

A Novel Function for Fragile X Mental Retardation Protein in Translational Activation

Fragile X syndrome, the most frequent form of inherited mental retardation, is due to the absence of Fragile X Mental Retardation Protein (FMRP), an RNA-binding protein involved in several steps of RNA metabolism. To date, two RNA motifs have been found to mediate FMRP/RNA interaction, the G-quartet and the “kissing complex,” which both induce translational repression in the presence of FMRP. We show here a new role for FMRP as a positive modulator of translation. FMRP specifically binds Superoxide Dismutase 1 (Sod1) mRNA with high affinity through a novel RNA motif, SoSLIP (Sod1 mRNA Stem Loops Interacting with FMRP), which is folded as three independent stem-loop structures. FMRP induces a structural modification of the SoSLIP motif upon its interaction with it. SoSLIP also behaves as a translational activator whose action is potentiated by the interaction with FMRP. The absence of FMRP results in decreased expression of Sod1. Because it has been observed that brain metabolism of FMR1 null mice is more sensitive to oxidative stress, we propose that the deregulation of Sod1 expression may be at the basis of several traits of the physiopathology of the Fragile X syndrome, such as anxiety, sleep troubles, and autism.     

A Practical and Novel Method to Extract Genomic DNA from Blood Collection Kits for Plasma Protein Preservation

Laboratory tests can be done on the cellular or fluid portions of the blood. The use of different blood collection tubes determines the portion of the blood that can be analyzed (whole blood, plasma or serum). Laboratories involved in studying the genetic basis of human disorders rely on anticoagulated whole blood collected in EDTA-containing vacutainer as the source of DNA for genetic / genomic analysis. Because most clinical laboratories perform biochemical, serologic and viral testing as a first step in phenotypic outcome investigation, anticoagulated blood is also collected in heparin-containing tube (plasma tube). Therefore when DNA and plasma are needed for simultaneous and parallel analyses of both genomic and proteomic data, it is customary to collect blood in both EDTA and heparin tubes. If blood could be collected in a single tube and serve as a source for both plasma and DNA, that method would be considered an advancement to existing methods. The use of the compacted blood after plasma extraction represents an alternative source for genomic DNA, thus minimizing the amount of blood samples processed and reducing the number of samples required from each patient. This would ultimately save time and resources.The BD P100 blood collection system for plasma protein preservation were created as an improved method over previous plasma or serum collection tubes¹, to stabilize the protein content of blood, enabling better protein biomarker discovery and proteomics experimentation from human blood. The BD P100 tubes contain 15.8 ml of spray-dried K2EDTA and a lyophilized proprietary broad spectrum cocktail of protease inhibitors to prevent coagulation and stabilize the plasma proteins. They also include a mechanical separator, which provides a physical barrier between plasma and cell pellets after centrifugation. Few methods have been devised to extract DNA from clotted blood samples collected in old plasma tubes^2-4. Challenges from these methods were mainly associated with the type of separator inside the tubes (gel separator) and included difficulty in recovering the clotted blood, the inconvenience of fragmenting or dispersing the clot, and obstruction of the clot extraction by the separation gel.We present the first method that extracts and purifies genomic DNA from blood drawn in the new BD P100 tubes. We compare the quality of the DNA sample from P100 tubes to that from EDTA tubes. Our approach is simple and efficient. It involves four major steps as follows: 1) the use of a plasma BD P100 (BD Diagnostics, Sparks, MD, USA) tube with mechanical separator for blood collection, 2) the removal of the mechanical separator using a combination of sucrose and a sterile paperclip metallic hook, 3) the separation of the buffy coat layer containing the white cells and 4) the isolation of the genomic DNA from the buffy coat using a regular commercial DNA extraction kit or a similar standard protocol.     

基于局部茎搜索的RNA二级结构预测算法

RNA的二级结构预测是生物信息学中一个已经有30多年历史的经典问题,基于最小自由能模型(MFE)的优化算法是使用最为广泛的方法．但RNA结构中假结的存在使MFE问题理论上成为一个NP-hard问题,即使采用动态规划等优化算法也会面临时间复杂度高的困难,同时研究还发现,由于受RNA折叠动力学机制以及环境因素的影响,真实的RNA二级结构往往并不处于自由能最小状态．根据RNA折叠的特点,提出了一种启发式搜索算法来预测带假结的RNA二级结构．该算法以RNA的茎为基本单元,采用启发式搜索策略在茎的组合空间中搜索自由能最小并且出现频率最高的RNA二级结构,该算法不仅能显著降低搜索RNA二级结构的时间复杂度,还有助于弥补单纯依赖能量预测RNA二级结构的不足．在多种类型的RNA标准数据集上进行了检验,结果表明,该算法在预测的精度上优于目前国际上几个著名的RNA二级结构预测算法并且具有较高的运行效率．     

A DEK Domain-Containing Protein Modulates Chromatin Structure and Function in Arabidopsis

Chromatin is a major determinant in the regulation of virtually all DNA-dependent processes. Chromatin architectural proteins interact with nucleosomes to modulate chromatin accessibility and higher-order chromatin structure. The evolutionarily conserved DEK domain-containing protein is implicated in important chromatin-related processes in animals, but little is known about its DNA targets and protein interaction partners. In plants, the role of DEK has remained elusive. In this work, we identified DEK3 as a chromatin-associated protein in Arabidopsis thaliana. DEK3 specifically binds histones H3 and H4. Purification of other proteins associated with nuclear DEK3 also established DNA topoisomerase 1α and proteins of the cohesion complex as in vivo interaction partners. Genome-wide mapping of DEK3 binding sites by chromatin immunoprecipitation followed by deep sequencing revealed enrichment of DEK3 at protein-coding genes throughout the genome. Using DEK3 knockout and overexpressor lines, we show that DEK3 affects nucleosome occupancy and chromatin accessibility and modulates the expression of DEK3 target genes. Furthermore, functional levels of DEK3 are crucial for stress tolerance. Overall, data indicate that DEK3 contributes to modulation of Arabidopsis chromatin structure and function.