DNA sequence analysis of a rat RT1 class 11 A β gene

The rat major histocompatibility complex (RT1) encodes twin sets of class II molecules, each consisting of two polypeptide chains referred to as A and A , and E and E . A gene encoding the RTLA chain was isolated from a rat genomic library using an HLA-DQ chain cDNA as a probe. The nucleotide sequence of the coding regions of this gene was determined. Comparison of this sequence with those of the corresponding genes of mouse (H-2A ) and human (HLA-DQ ) revealed that this gene has been highly conserved during evolution, and that some parts of the molecule are more conserved than others. Analysis of the nucleotide sequence encoding the two external domains suggests that the membrane proximal domain has been subject to conservative selection, whereas replacement substitutions have been selected positively at certain residues within the amino terminal domain. The overall organization of the RT1.A gene is similar to that of the H-2A gene.     

AliGROOVE – visualization of heterogeneous sequence divergence within multiple sequence alignments and detection of inflated branch support

Background

Masking of multiple sequence alignment blocks has become a powerful method to enhance the tree-likeness of the underlying data. However, existing masking approaches are insensitive to heterogeneous sequence divergence which can mislead tree reconstructions. We present AliGROOVE, a new method based on a sliding window and a Monte Carlo resampling approach, that visualizes heterogeneous sequence divergence or alignment ambiguity related to single taxa or subsets of taxa within a multiple sequence alignment and tags suspicious branches on a given tree.

Results

We used simulated multiple sequence alignments to show that the extent of alignment ambiguity in pairwise sequence comparison is correlated with the frequency of misplaced taxa in tree reconstructions. The approach implemented in AliGROOVE allows to detect nodes within a tree that are supported despite the absence of phylogenetic signal in the underlying multiple sequence alignment. We show that AliGROOVE equally well detects heterogeneous sequence divergence in a case study based on an empirical data set of mitochondrial DNA sequences of chelicerates.

Conclusions

The AliGROOVE approach has the potential to identify single taxa or subsets of taxa which show predominantly randomized sequence similarity in comparison with other taxa in a multiple sequence alignment. It further allows to evaluate the reliability of node support in a novel way.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2105-15-294) contains supplementary material, which is available to authorized users.     

The limits of protein sequence comparison?

Modern sequence alignment algorithms are used routinely to identify homologous proteins, proteins that share a common ancestor. Homologous proteins always share similar structures and often have similar functions. Over the past 20 years, sequence comparison has become both more sensitive, largely because of profile-based methods, and more reliable, because of more accurate statistical estimates. As sequence and structure databases become larger, and comparison methods become more powerful, reliable statistical estimates will become even more important for distinguishing similarities that are due to homology from those that are due to analogy (convergence). The newest sequence alignment methods are more sensitive than older methods, but more accurate statistical estimates are needed for their full power to be realized.     

The nucleotide sequence of bacteriophage φX174

The complete nucleotide sequence of the DNA of bacteriophage φX174 has been determined. The provisional sequence (Sanger et al., 1977a) deduced largely by the plus and minus method, has been completed and confirmed, predominantly using the terminator method (Sanger et al., 1977b). About 30 revisions were found to be necessary in the 5386-nucleotide sequence. The amino acid sequences of the ten proteins for which the DNA codes have also been deduced.     

What determines the rate of sequence evolution?

High rates of amino-acid sequence evolution have sometimes been considered to be diagnostic for genes undergoing adaptive change. However, two recent studies have shown that rapid evolution of amino-acid sequence can also be congruent with neutrality.     

Isolation and partial sequence of goat spleen prothymosin α

Goat prothymosin , a highly acidic polypeptide of pl 3.5, 109 amino acid residues, has been isolated from lymphoid and non-lymphoid tissues of young female goats. Unlike rat, murine and porcine prothymosins , goat prothymosin appears at a higher concentration in the spleen compared with the thymus. The sequence of segments of the polypeptide involving known mutations has been determined, by automatic sequencing of its tryptic peptide fragments. The acidic amino acid-rich segment in the middle of the molecule, including residues 49–83, has not been sequenced. Goat prothymosin closely resembles bovine prothymosin , with only one substitution, proline for alanine at position 85. It also resembles human prothymosin , with only three substitutions. It differs more significantly from rat and murine prothymosins , by two deletions and three substitutions. The results show the highly conserved nature of the molecule, with substitutions at given positions only.Abbreviations ProT Prothymosin - T₁ Thymosin ₁ - MLR Mixed Lymphocyte Response - HPLC High Performance Liquid Chromatography - RIA Radioimmunoassay - B Aspartic acid or Asparagine - Z Glutamic acid or Glutamine     

The sequence upstream of the −10 consensus sequence modulates the strength and induction time of stationary-phase promoters in Escherichia coli

We constructed a library of synthetic stationary-phase promoters for Escherichia coli. For designing the promoters, the known −10 consensus sequence, as well as the extended −10 region, and an A/T-rich region downstream of the −10 region were kept constant, whereas sequences from −37 to −14 were partially or completely randomised. For detection and selection of stationary-phase promoters, green fluorescent protein (GFP) with enhanced fluorescence was used. To establish the library, 33 promoters were selected, which differ in strength from 670 to more than 13,000 specific fluorescence units, indicating that the strength of promoters can be modulated by the sequence upstream of the −10 region. DNA sequencing revealed a preferential insertion of nucleotides depending on the position. By expressing the promoters in an rpoS-deficient strain, a special group of stationary-phase promoters was identified, which were expressed exclusively or preferentially by RNA polymerase holoenzyme Eσ^s. The DNA sequence of these promoters differed significantly in the region from −25 to −16. Furthermore, it was shown that the DNA curvature of the promoter region had no effect on promoter strength. The broad range of promoter activities make these promoters very suitable for fine-tuning of gene expression and for cost-effective large-scale applications in industrial bioprocesses.     

Isolation and characterization of the mitochondrial ATP synthase fromChlamydomonas reinhardtii. cDNA sequence and deduced protein sequence of the α subunit

We have isolated the F₀F₁-ATP synthase complex from oligomycin-sensitive mitochondria of the green algaChlamydomonas reinhardtii. A pure and active ATP synthase was obtained by eans of sonication, extraction with dodecyl maltoside and ion exchange and gel permeation chromatography in the presence of glycerol, DTT, ATP and-21. The enzyme consists of 14 subunits as judged by SDS-PAGE. A cDNA clone encoding the ATP synthase subunit has been sequenced. The deduced protein sequence contains a presequence of 45 amino acids which is not present in the mature protein. The mature protein is 58–70% identical to corresponding mitochondrial proteins from other organisms. In contrast to the ATP synthase subunit fromC. reinhardtii (Franzen and Falk, Plant Mol Biol 19 (1992) 771–780), the protein does not have a C-terminal extension. However, the N-terminal domain of the mature protein is 15–18 residues longer than in ATP synthase subunits from other organisms. Southern blot analysis indicates that the protein is encoded by a single-copy gene.Abbreviations DM dodecyl--D-maltoside - OSCP oligomycin sensitivity conferring protein - PMSF phenyl-methylsulfonylfluoride - DTT dithiothreitol - EDTA ethylenediaminotetraacetic disodium salt     

Nucleotide sequence of the humanθ 1-globin gene

We have cloned and sequenced the human ₁-globin gene. The nucleotide sequence and organization of the human ₁ gene (exons, introns, promoter, and polyadenylation signals) are similar to those reported for the orangutan ₁-globin gene. If these genes are functional, the sequences of their ₁-globin chains would differ by only one amino acid residue (at position 137).This research was supported by USPHS Research Grants HLB-05168 and HLB-15158. This is contribution No. 1085 from the Department of Cell and Molecular Biology at the Medical College of Georgia in Augusta.     

5′ Flanking sequence of the hamster desmin gene

Molecular Biology Reports -     

A reevaluation of the amino acid sequence of human follitropin β-subunit

A collaborative study from two laboratories has been undertaken to re-evaluate the human follitropin β-subunit sequence (hFSHβ), since areas of uncertainty remain in the wake of two earlier reports. The first report was by Shome and Parlow (1974). The second, by Saxena and Rathnam (1976), proposed revisions for sequence not definitively placed in the first study, as well as some differences in other placements. We have re-examined the sequence of the hFSHβ with more recent methodology. This has led to revision of certain areas of the sequence and resolution of differences between the two earlier proposals. Specifically, an-Ile-Ser- is established at 21–22, Asp at 41, Arg at 44, Lys at 46, and Glu at 111. These were areas of disagreement in the earlier proposals. A definitive placement of the residues around tryptophan-27 has now been obtained by three laboratories. C-terminal heterogeneity was observed with subunits ending at residue 107, 109, or 111. N-terminal heterogeneity has been observed in all preparations examined to date. A significant population of molecules with a proteolytic nick between residues 38–39 is noted. This is very likely an artifact of the collection and processing. The preparations examined in the present studies showed no evidence of residues 112–118 proposed by Saxena and Rathnam.     

Evolution: What determines the rate of sequence evolution?

High rates of amino-acid sequence evolution have sometimes been considered to be diagnostic for genes undergoing adaptive change. However, two recent studies have shown that rapid evolution of amino-acid sequence can also be congruent with neutrality.     

A polymorphic poly-A sequence in the 5′ region of the aldosynthase (CYP11B2) gene may be useful in genetic diagnosis of 11β-hydroxylase genes defects

We report a highly polymorphic poly-A sequence in the 5 flanking region of CYP11B2 that may be used in genetic counselling for prenatal diagnosis or carrier detection in families affected by steroid 11-hydroxylase genes defects.     

Quantifying sequence and structural features of protein–RNA interactions

Increasing awareness of the importance of protein–RNA interactions has motivated many approaches to predict residue-level RNA binding sites in proteins based on sequence or structural characteristics. Sequence-based predictors are usually high in sensitivity but low in specificity; conversely structure-based predictors tend to have high specificity, but lower sensitivity. Here we quantified the contribution of both sequence- and structure-based features as indicators of RNA-binding propensity using a machine-learning approach. In order to capture structural information for proteins without a known structure, we used homology modeling to extract the relevant structural features. Several novel and modified features enhanced the accuracy of residue-level RNA-binding propensity beyond what has been reported previously, including by meta-prediction servers. These features include: hidden Markov model-based evolutionary conservation, surface deformations based on the Laplacian norm formalism, and relative solvent accessibility partitioned into backbone and side chain contributions. We constructed a web server called aaRNA that implements the proposed method and demonstrate its use in identifying putative RNA binding sites.     

cDNA sequence and chromosome localization of pig α1,3 galactosyltransferase

Human serum contains natural antibodies (NAb), which can bind to endothelial cell surface antigens of other mammals. This is believed to be the major initiating event in the process of hyperacute rejection of pig to primate xenografts. Recent work has implicated galoctosyl 1,3 galactosyl 1,4 N-acetyl-glucosaminyl carbohydrate epitopes, on the surface of pig endothelial cells as a major target of human natural antibodies. This epitope is made by a specific galactosyltransferase (1,3 GT) present in pigs but not in higher primates. We have now cloned and sequenced a full-length pig 1,3 GT cDNA. The predicted 371 amino acid protein sequence shares 85% and 76% identity with previously characterized cattle and mouse 1,3 GT protein sequences, respectively. By using fluorescence and isotopic in situ hybridization, the GGTA1 gene was mapped to the region q2.10–q2.11 of pig chromosome 1, providing further evidence of homology between the subterminal region of pig chromosome 1q and human chromosome 9q, which harbors the locus encoding the AB0 blood group system, as well as a human pseudogene homologous to the pig GGTA1 gene.The nucleotide sequence data reported in this paper have been submitted to the GenBank nucleotide sequence database and have been assigned the accession number L36152     

Locust solid cuticle—A time sequence of mechanical properties

Studies on locust femur solid cuticle indicate that there is a continuum of biomechanically important properties from one instar to the next, and that there is a constant ratio of stiffness to mass which is only interrupted at ecdysis. The biochemical nature of both larval and adult solid cuticle is interpreted in terms of both the tangent modulus and the breaking strength of the femur cuticle. The mechanical behaviour of the different layers in the cuticle is discussed.     

Characterization and sequence prediction of structural variations in α-helix

Background

Centralised resources such as GenBank and UniProt are perfect examples of the major international efforts that have been made to integrate and share biological information. However, additional data that adds value to these resources needs a simple and rapid route to public access. The Distributed Annotation System (DAS) provides an adequate environment to integrate genomic and proteomic information from multiple sources, making this information accessible to the community. DAS offers a way to distribute and access information but it does not provide domain experts with the mechanisms to participate in the curation process of the available biological entities and their annotations.

Results

We designed and developed a Collaborative Annotation System for proteins called DAS Writeback. DAS writeback is a protocol extension of DAS to provide the functionalities of adding, editing and deleting annotations. We implemented this new specification as extensions of both a DAS server and a DAS client. The architecture was designed with the involvement of the DAS community and it was improved after performing usability experiments emulating a real annotation task.

Conclusions

We demonstrate that DAS Writeback is effective, usable and will provide the appropriate environment for the creation and evolution of community protein annotation.