Alignment-free Comparison of Protein Sequences Based on Reduced Amino Acid Alphabets

Abstract

Protein sequences are treated as stochastic processes on the basis of a reduced amino acid alphabet of 10 types of amino acids. The realization of a stochastic process is described by associated transition probability matrix that corresponds to the process uniquely. Then new distances between transition probability matrices are defined for sequences similarity analysis. Two separate datasets are prepared and tested to identify the validity of the method. The results demonstrate the new method is powerful and efficient.     

Amino Acid Availability Regulates the Effect of Hyperinsulinemia on Skin Protein Metabolism in Pigs

The effects of amino acid supply and insulin infusion on skin protein kinetics (fractional synthesis rate (FSR), fractional breakdown rate (FBR), and net balance (NB)) in pigs were investigated. Four-month-old pigs were divided into four groups as follows: control, insulin (INS), amino acid (AA), and INS + AA groups based on the nutritional and hormonal conditions. l-[ring-¹³C₆]Phenylalanine was infused. FBR was estimated from the enrichment ratio of arterial phenylalanine to intracellular free phenylalanine. Plasma INS was increased (p < 0.05) in the INS and INS + AA groups. Plasma glucose was maintained by infusion of glucose in the groups receiving INS. The interventions did not change the NB of skin protein. However, the interventions affected the FSR and FBR differently. An infusion of INS significantly increased both FSR and FBR, although AA infusion did not. When an AA infusion was added to the infusion of insulin (INS + AA group), FSR and FBR were both lower when compared with the INS group. Our data demonstrate that in anesthetized pigs INS infusion did not exert an anabolic effect, but rather it increased AA cycling into and out of skin protein. Because co-infusion of AAs with INS ameliorated this effect, it is likely that the increased AA cycling during INS infusion was related to AA supply. Although protein kinetics were affected by both INS and AAs, none of the interventions affected the skin protein deposition. Thus, skin protein content is closely regulated under normal circumstances and is not subject to transient changes in AAs or hormonal concentrations.     

Amino Acid Sequences of Glycopeptides from Cucumisin

Oxidation of short-chain iso-alkanes (isobutane, isopentane, 2-methylpentane, and 3-methylpentane) was studied with propane-grown resting mycelia of Scedosporium sp. A-4. Isobutane was oxidized to terf-butanol, but both isobutane and tert-butanol were not used for growth. Isopentane was oxidized to 3-methyl-1-butanol, 2-methyl-2-butanol, and 3-methyl-2-butanol but not to 2-methyl-1-butanol. 2-Methylpentane was oxidized to 4-methyl-1-pentanol, 2-methyl-2-pentanol, and 4-methyl-2-pentanol but not to 2-methyl-1-pentanol or 2-methyl-3-pentanol. 3-Methylpentane was not oxidized. Oxidation of branched alcohols was also studied.     

Human Protein Cluster Analysis Using Amino Acid Frequencies

The paper focuses on the development of a software tool for protein clustering according to their amino acid content. All known human proteins were clustered according to the relative frequencies of their amino acids starting from the UniProtKB/Swiss-Prot reference database and making use of hierarchical cluster analysis. Results were compared to those based on sequence similarities. Results: Proteins display different clustering patterns according to type. Many extracellular proteins with highly specific and repetitive sequences (keratins, collagens etc.) cluster clearly confirming the accuracy of the clustering method. In our case clustering by sequence and amino acid content overlaps. Proteins with a more complex structure with multiple domains (catalytic, extracellular, transmembrane etc.), even if classified very similar according to sequence similarity and function (aquaporins, cadherins, steroid 5-alpha reductase etc.) showed different clustering according to amino acid content. Availability of essential amino acids according to local conditions (starvation, low or high oxygen, cell cycle phase etc.) may be a limiting factor in protein synthesis, whatever the mRNA level. This type of protein clustering may therefore prove a valuable tool in identifying so far unknown metabolic connections and constraints.     

A Combinatorial Approach to Detect Coevolved Amino Acid Networks in Protein Families of Variable Divergence

Communication between distant sites often defines the biological role of a protein: amino acid long-range interactions are as important in binding specificity, allosteric regulation and conformational change as residues directly contacting the substrate. The maintaining of functional and structural coupling of long-range interacting residues requires coevolution of these residues. Networks of interaction between coevolved residues can be reconstructed, and from the networks, one can possibly derive insights into functional mechanisms for the protein family. We propose a combinatorial method for mapping conserved networks of amino acid interactions in a protein which is based on the analysis of a set of aligned sequences, the associated distance tree and the combinatorics of its subtrees. The degree of coevolution of all pairs of coevolved residues is identified numerically, and networks are reconstructed with a dedicated clustering algorithm. The method drops the constraints on high sequence divergence limiting the range of applicability of the statistical approaches previously proposed. We apply the method to four protein families where we show an accurate detection of functional networks and the possibility to treat sets of protein sequences of variable divergence.     

CodonTest: Modeling Amino Acid Substitution Preferences in Coding Sequences

Codon models of evolution have facilitated the interpretation of selective forces operating on genomes. These models, however, assume a single rate of non-synonymous substitution irrespective of the nature of amino acids being exchanged. Recent developments have shown that models which allow for amino acid pairs to have independent rates of substitution offer improved fit over single rate models. However, these approaches have been limited by the necessity for large alignments in their estimation. An alternative approach is to assume that substitution rates between amino acid pairs can be subdivided into rate classes, dependent on the information content of the alignment. However, given the combinatorially large number of such models, an efficient model search strategy is needed. Here we develop a Genetic Algorithm (GA) method for the estimation of such models. A GA is used to assign amino acid substitution pairs to a series of rate classes, where is estimated from the alignment. Other parameters of the phylogenetic Markov model, including substitution rates, character frequencies and branch lengths are estimated using standard maximum likelihood optimization procedures. We apply the GA to empirical alignments and show improved model fit over existing models of codon evolution. Our results suggest that current models are poor approximations of protein evolution and thus gene and organism specific multi-rate models that incorporate amino acid substitution biases are preferred. We further anticipate that the clustering of amino acid substitution rates into classes will be biologically informative, such that genes with similar functions exhibit similar clustering, and hence this clustering will be useful for the evolutionary fingerprinting of genes.     

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     

A New Method of Inference of Ancestral Nucleotide and Amino Acid Sequences

A statistical method was developed for reconstructing the nucleotide or amino acid sequences of extinct ancestors, given the phylogeny and sequences of the extant species. A model of nucleotide or amino acid substitution was employed to analyze data of the present-day sequences, and maximum likelihood estimates of parameters such as branch lengths were used to compare the posterior probabilities of assignments of character states (nucleotides or amino acids) to interior nodes of the tree; the assignment having the highest probability was the best reconstruction at the site. The lysozyme c sequences of six mammals were analyzed by using the likelihood and parsimony methods. The new likelihood-based method was found to be superior to the parsimony method. The probability that the amino acids for all interior nodes at a site reconstructed by the new method are correct was calculated to be 0.91, 0.86, and 0.73 for all, variable, and parsimony-informative sites, respectively, whereas the corresponding probabilities for the parsimony method were 0.84, 0.76, and 0.51, respectively. The probability that an amino acid in an ancestral sequence is correctly reconstructed by the likelihood analysis ranged from 91.3 to 98.7% for the four ancestral sequences.     

基于进化信息改进蛋白质定点突变稳定性预测准确率

定点突变后蛋白质稳定性的增加还是降低,是分子生物学和蛋白质工程的核心问题之一,也是目前生物信息学研究的重要领域。基于蛋白质序列信息对蛋白质定点突变后的稳定性进行预测的方法,因其简易、适用面广而得到广泛的研究应用。通过对编码策略（coding schemes）的探索,发现不同编码策略对预测准确率有较大影响,并发现基于进化信息的BLOSUM打分矩阵可以用于蛋白质定点突变稳定性预测,具有较高的预测准确率。应用基于BLOSUM62打分矩阵的神经网络（ANN）和支持向量机（SVM）算法,可以改进蛋白质定点突变后稳定性的预测,而且ANN+ BLOSUM62在1623条序列的数据集上的实测结果优于目前国际通用的几款预测 软件。     

Evolution and Taxonomy of Positive-Strand RNA Viruses: Implications of Comparative Analysis of Amino Acid Sequences

Abstract

Despite the rapid mutational change that is typical of positive-strand RNA viruses, enzymes mediating the replication and expression of virus genomes contain arrays of conserved sequence motifs. Proteins with such motifs include RNA-dependent RNA polymerase, putative RNA helicase, chymotrypsin-like and papain-like proteases, and methyltransferases. The genes for these proteins form partially conserved modules in large subsets of viruses. A concept of the virus genome as a relatively evolutionarily stable “core” of housekeeping genes accompanied by a much more flexible “shell” consisting mostly of genes coding for virion components and various accessory proteins is discussed. Shuffling of the “shell” genes including genome reorganization and recombination between remote groups of viruses is considered to be one of the major factors of virus evolution.

Multiple alignments for the conserved viral proteins were constructed and used to generate the respective phylogenetic trees. Based primarily on the tentative phylogeny for the RNA-dependent RNA polymerase, which is the only universally conserved protein of positive-strand RNA viruses, three large classes of viruses, each consisting of distinct smaller divisions, were delineated. A strong correlation was observed between this grouping and the tentative phylogenies for the other conserved proteins as well as the arrangement of genes encoding these proteins in the virus genome. A comparable correlation with the polymerase phylogeny was not found for genes encoding virion components or for genome expression strategies. It is surmised that several types of arrangement of the “shell” genes as well as basic mechanisms of expression could have evolved independently in different evolutionary lineages.

The grouping revealed by phylogenetic analysis may provide the basis for revision of virus classification, and phylogenetic taxonomy of positive-strand RNA viruses is outlined. Some of the phylogenetically derived divisions of positive-strand RNA viruses also include double-stranded RNA viruses, indicating that in certain cases the type of genome nucleic acid may not be a reliable taxonomic criterion for viruses.

Hypothetical evolutionary scenarios for positive-strand RNA viruses are proposed. It is hypothesized that all positive-strand RNA viruses and some related double-stranded RNA viruses could have evolved from a common ancestor virus that contained genes for RNA-dependent RNA polymerase, a chymotrypsin-related protease that also functioned as the capsid protein, and possibly an RNA helicase.     

基于编码序列、基因间序列和氨基酸序列构建的系统发生关系比较

以7种古菌、46种细菌和10种真核生物的基因组为样本,考虑碱基间的短程关联和长程关联作用,得到编码序列的密码对和基因间序列的三联体对中不同位点的二核苷酸频率,据此构建了基于编码序列和基因间序列的系统发生关系。无论是基于编码序列还是基因间序列对信息进行聚类,古菌或真核均被聚在一支上,表明聚类参数的选择是合适的;与基于氨基酸序列构建的系统发生关系进行两两比较,发现大部分硬壁菌的编码序列与基因间序列之间,以及编码序列与氨基酸序列之间的进化都存在较大差异。通过分析认为,只有综合考虑这三类序列的进化信息,才可能得到更自然的系统发生关系。     

Effect of Anaplerotic Fluxes and Amino Acid Availability on Hepatic Lipoapoptosis

To identify metabolic pathways involved in hepatic lipoapoptosis, metabolic flux analysis using [U-¹³C₅]glutamine as an isotopic tracer was applied to quantify phenotypic changes in H4IIEC3 hepatoma cells treated with either palmitate alone (PA-cells) or both palmitate and oleate in combination (PA/OA-cells). Our results indicate that palmitate inhibited glycolysis and lactate dehydrogenase fluxes while activating citric acid cycle (CAC) flux and glutamine uptake. This decoupling of glycolysis and CAC fluxes occurred during the period following palmitate exposure but preceding the onset of apoptosis. Oleate co-treatment restored most fluxes to their control levels, resulting in steatotic lipid accumulation while preventing apoptosis. In addition, palmitate strongly increased the cytosolic NAD⁺/NADH ratio, whereas oleate co-treatment had the opposite effect on cellular redox. We next examined the influence of amino acids on these free fatty acid-induced phenotypic changes. Increased medium amino acids enhanced reactive oxygen species (ROS) generation and apoptosis in PA-cells but not in PA/OA-cells. Overloading the medium with non-essential amino acids induced apoptosis, but essential amino acid overloading partially ameliorated apoptosis. Glutamate was the most effective single amino acid in promoting ROS. Amino acid overloading also increased cellular palmitoyl-ceramide; however, ceramide synthesis inhibitors had no effect on measurable indicators of apoptosis. Our results indicate that free fatty acid-induced ROS generation and apoptosis are accompanied by the decoupling of glycolysis and CAC fluxes leading to abnormal cytosolic redox states. Amino acids play a modulatory role in these processes via a mechanism that does not involve ceramide accumulation.     

SuccSite: Incorporating Amino Acid Composition and Informative k-spaced Amino Acid Pairs to Identify Protein Succinylation Sites

Protein succinylation is a biochemical reaction in which a succinyl group (-CO-CH2-CH2-CO-) is attached to the lysine residue of a protein molecule. Lysine succinylation plays important regulatory roles in living cells. However, studies in this field are limited by the difficulty in experimentally identifying the substrate site specificity of lysine succinylation. To facilitate this process, several tools have been proposed for the computational identification of succinylated lysine sites. In this study, we developed an approach to investigate the substrate specificity of lysine succinylated sites based on amino acid composition. Using experimentally verified lysine succinylated sites collected from public resources, the significant differences in position-specific amino acid composition between succinylated and non-succinylated sites were represented using the Two Sample Logo program. These findings enabled the adoption of an effective machine learning method, support vector machine, to train a predictive model with not only the amino acid composition, but also the composition of k-spaced amino acid pairs. After the selection of the best model using a ten-fold cross-validation approach, the selected model significantly outperformed existing tools based on an independent dataset manually extracted from published research articles. Finally, the selected model was used to develop a web-based tool, SuccSite, to aid the study of protein succinylation. Two proteins were used as case studies on the website to demonstrate the effective prediction of succinylation sites. We will regularly update SuccSite by integrating more experimental datasets. SuccSite is freely accessible at http://csb.cse.yzu.edu.tw/SuccSite/.     

Rice Embryo Globulins: Amino-Terminal Amino Acid Sequences, cDNA Cloning and Expression

A globulin fraction prepared from rice embryos contained polypeptidesor polypeptide groups of 49 kDa (designated REG1), 46 kDa (designatedREG2), about 35 kDa, 32 kDa and 25 kDa. The amino-terminal sequencesof REG1 and the major polypeptide in the 35-kDa group were identical,suggesting that the REG1 polypeptide undergoes partial proteolyticprocessing that removes a carboxy-terminal region. A cDNA clone,designated pcREG2, encoding REG2 was isolated, and its nucleotidesequence was determined. The deduced amino acid sequence ofREG2 was found to be 68% identical to that of the maize GLB2globulin. Reg2 mRNA was present at high levels during embryodevelopment for up to 14 days after flowering (DAF). Lower levelswere found 20 DAF when the maturation of embryos was almostcompleted, and at the dry mature stage. Reg2 mRNA almost disappearedupon imbibition of isolated dry mature embryos but it was re-inducedat a low level by further treatment with ABA. The expressionof Reg2 was not induced by ABA in suspension-cultured cells,unlike that of Osem, one of the late embryogenesis abundantprotein (LEA) genes. (Received November 6, 1995; Accepted April 22, 1996)     

Amino Acid Sequences of Porcine Fast and Slow Troponin T Isoforms

In this study, 10 troponin T isoforms from adult porcine skeletal muscle messenger RNA were clarified. These were eight fast- and two slow-type isoforms. Fast-type isoforms had three and two variable exons in the N-terminal and the C-terminal region respectively. Slow-type isoforms had one variable exon in the N-terminal region.     

Conservation of Amino Acid Sequences Between Human and Porcine Choline Acetyltransferase

Abstract: The amino acid sequence of 11 peptides generated from human placental choline acetyltransferase was compared to the corresponding amino acid sequences predicted from the nucleotide sequence of a recently cloned porcine choline acetyltransferase cDNA. These peptides, which were generated by cyanogen bromide cleavage or tryptic digestion, accounted for 23% of the amino acids in the enzyme. Of the 145 amino acids sequenced eight differed between the two species, yielding an identity of 94% over the regions sampled.
Of the eight amino acids that differed six could represent single base changes in the DNA sequence. These findings demonstrate strong sequence similarity between porcine and human choline acetyltransferase and indicate that they are closely related evolutionarily.     

氨基酸组成对蛋白质耐热性的影响

为了研究一级结构对蛋白质耐热性的影响,利用软件DNAMAN对16个家族32种蛋白质序列进行了氨基酸含量分析,并统计分析了氨基酸组成对蛋白质耐热性的影响。通过比较同一家族的高低温蛋白质序列及16个家族中所有高温和低温蛋白质序列中氨基酸含量的变化可以推断(从低温到高温)：Ser、Cys．含量降低显著,Arg、Ile、Pro含量升高显著。由此可知高温蛋白质倾向于含有疏水性氨基酸而避免亲水性氨基酸。     

Amino Acid Sequence of Porcine Myelin Basic Protein

The myelin basic protein (BP) of pig brain was cleaved into its constituent tryptic peptides and the amino acid composition of each was determined. Those tryptic peptides that had not been sequenced previously were cleaved with dipeptidyl peptidases and the resulting dipeptides were trimethylsilated, separated by gas chromatography, and identified by mass spectrometry. Carboxypeptidases B and Y were used to establish the COOH-terminal sequences of some of the tryptic peptides; one tryptic peptide (sequence 76-92) was cleaved with thermolysin and the thermolytic peptides were analyzed. From the results of the present study together with those reported previously, it has been possible to determine the complete amino acid sequence of the protein. The protein consists of 172 residues and has a theoretical molecular weight of 18,604. Its amino acid sequence is identical with that reported for the homologous bovine protein with the following exceptions: Ser replaces (bovine) Ala2; His-Gly is inserted between Arg9 and Ser10; Ala replaces Ser45; His and Gly replace Gly76 and His77, respectively; Pro replaces Ser131 and Ser135; Ala is inserted between Gly142 and His143; and Gln replaces His143.     

氨基酸在蛋白质空间结构中的深度倾向性因子

用统计和几何方法给出了氨基酸在蛋白质空间结构中的深度计算,并利用PDB数据库得到了不同氨基酸在蛋白质中的深度倾向性因子,并得到了这些倾向性因子与氨基酸的物理、化学综合特性的相关性质.